ArticlePDF Available

Glutathione transferases in bacteria

October 2008

October 2008
276:58-75

Authors:

Nerino Allocati

University “G. d' Annunzio” of Chieti - Pescara

Luca Federici

Università degli Studi G. d'Annunzio Chieti e Pescara

Structural studies on beta class GSTs. (A) Structure of OaGST shown with the twofold symmetry axis perpendicular to the page (pdb code: 2NTO). The N-terminal thioredoxin domain and the C-terminal all helical domain are highlighted with different tonalities. GSH molecules are shown in sticks. (B) Overlay of four different beta class GSTs dimers shown as Ca traces: Oa-GST (red), BxGST (green, pdb code: 2GDR), EcGST (magenta, pdb code: 1A0F) and PmGST (cyan, pdb code: 1PMT). This representation highlights the conservation of beta class fold as well as of the monomers' orientation in the dimers. (C) GSH binding site in PmGST. In this crystal structure, GSH (green carbons) forms a mixed disulfide with Cys10. His106 and Ser9 are also at hydrogen bond distance from the GSH sulfur. (D) Structure of BxGST in complex with two GSH molecules (green carbons), one at the canonical G-site and the other bound at the hydrophobic substrates binding site (H-site). The two GSH sulfurs are at close distance from each other. This figure was prepared with PYMOL (DeLano Scientific).

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REVIEW ARTICLE

Glutathione transferases in bacteria

Nerino Allocati

, Luca Federici

1,2

, Michele Masulli

and Carmine Di Ilio

1,2

1 Dipartimento di Scienze Biomediche, Universita

`‘G. d’Annunzio’, Chieti, Italy

2 Ce.S.I. Centro Studi per l’Invecchiamento, Universita

`‘G. d’Annunzio’, Chieti, Italy

Introduction

Glutathione transferases (GSTs; EC 2.5.1.18) consti-

tute a protein superfamily that is involved in cellular

detoxiﬁcation against harmful xenobiotics and endo-

biotics [1–3]. One common feature of all GSTs is their

ability to catalyse nucleophilic attack by the tripeptide

glutathione (GSH) on the electrophilic groups of a

wide range of hydrophobic toxic compounds, thus pro-

moting their excretion from the cell [1]. GSTs are also

involved in several other cell functions, and are capa-

ble of binding a large number of endogenous and

exogenous compounds non-catalytically [1]. GSTs are

widely distributed in nature and are found in both

eukaryotes and prokaryotes. GSTs are divided into

at least four major families of proteins, namely cyto-

solic GSTs, mitochondrial GSTs, microsomal GSTs

and bacterial fosfomycin-resistance proteins [1,4]. The

cytosolic GSTs (cGSTs) have been subgrouped into

numerous divergent classes on the basis of their chemi-

cal, physical and structural properties [1,2]. The mito-

chondrial GSTs, also known as kappa class GSTs, are

soluble enzymes that have been characterized in

eukaryotes [5]. The third GST family comprises mem-

brane-bound transferases called membrane-associated

proteins involved in ecosanoid and glutathione metab-

olism (MAPEG), but these bear no similarity to solu-

ble GSTs [6]. Representatives of all three families are

also present in prokaryotes. The fourth family is found

exclusively in bacteria.

Keywords

bacterial glutathione transferase; chemical

stress; DCM dehalogenase; detoxiﬁcation;

drug resistance; fosfomycin resistance

proteins; glutathione; HCCA isomerase;

oxidative stress; TCHQ dehalogenase

Correspondence

N. Allocati, Dipartimento di Scienze

Biomediche, Universita’ ‘G. d’Annunzio’, Via

dei Vestini 31, I-66013 Chieti, Italy

Fax: +39 0871 355 5282

Tel: +39 0871 355 5281

E-mail: allocati@unich.it

(Received 25 July 2008, revised 8 October

2008, accepted 14 October 2008)

doi:10.1111/j.1742-4658.2008.06743.x

Bacterial glutathione transferases (GSTs) are part of a superfamily of

enzymes that play a key role in cellular detoxiﬁcation. GSTs are widely dis-

tributed in prokaryotes and are grouped into several classes. Bacterial

GSTs are implicated in a variety of distinct processes such as the biodegra-

dation of xenobiotics, protection against chemical and oxidative stresses

and antimicrobial drug resistance. In addition to their role in detoxiﬁca-

tion, bacterial GSTs are also involved in a variety of distinct metabolic

processes such as the biotransformation of dichloromethane, the degrada-

tion of lignin and atrazine, and the reductive dechlorination of pentachloro-

phenol. This review article summarizes the current status of knowledge

regarding the functional and structural properties of bacterial GSTs.

Abbreviations

BxGST, Burkholderia xenovorans GST; CDNB, 1-chloro-2,4-dinitrobenzene; cGST, cytosolic GST; DCM, dichloromethane; EcGST,

Escherichia coli GST; GSH, glutathione; GST, glutathione transferase; HCCA, 2-hydroxychromene-2-carboxylic acid; MAPEG, membrane-

associated proteins involved in ecosanoid and glutathione metabolism; OaGST, Ochrobactrum anthropi GST; PmGST, Proteus mirabilis GST;

TCHQ, tetrachlorohydroquinone.

58 FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS

The ﬁrst evidence for the presence of GSTs in bacteria

was reported more 25 years ago by Takashi Shishido

who showed the presence of GST activity in a strain of

Escherichia coli [7]. Since then, GSTs have been found

to be broadly distributed in aerobic prokaryotes, includ-

ing human and plant pathogens and soil bacteria. Con-

versely, GSTs have not been identiﬁed to date in

anaerobic bacteria [8] or in Archaea, the other domain

of prokaryotes. The absence of the enzyme in these

microorganisms is consistent with the lack of GSH [9].

This review summarizes current knowledge on the

functional and structural properties of bacterial GSTs

and the potential biotechnological applications of these

enzymes.

Classiﬁcation and phylogenetic

relationships

cGSTs, also termed canonical GSTs, are homo- or hete-

rodimeric enzymes found in aerobic forms of life. cGSTs

metabolize a broad range of electrophilic substrates via

GSH conjugation. They are involved in protecting cells

against oxidative stresses, have peroxidase and isomer-

ase activities and are implicated in the development of

drug resistance [1,2]. In mammalian species they are well

characterized and have been grouped into seven classes:

alpha, mu, pi, sigma, theta, omega and zeta [1]. Several

other classes are restricted to non-mammalian species:

beta, chi, delta, epsilon, lambda, phi and tau [1,2,10]. To

assign a cGST to a class, it is generally accepted that

proteins with > 40% sequence identity belong to the

same class, whereas GSTs of different classes share

< 25% sequence identity. The identity increases if the

N-terminal region only is considered. This region com-

prises part of the active site, with residues that interact

with GSH, and it is evolutionarily conserved [1,2,11].

Besides amino acid sequence identity, immunological

properties, kinetic features as well as similarity of the

crystal structures provide additional supporting data

[1,2].

In bacteria, four different classes of cGSTs have

been identiﬁed: beta, chi, theta and zeta [2,10,12,13].

Beta class cGSTs have been puriﬁed and characterized

from several bacteria [14–25]. They are able to conju-

gate the model substrate 1-chloro-2,4-dinitrobenzene

(CDNB) and bind to the GSH–afﬁnity matrix. All beta

class enzymes are characterized by the presence of a

cysteine residue at the GSH site [12]. The ﬁrst cGST of

this class was characterized from a Proteus mirabilis

strain (PmGST). PmGST displayed biochemical and

structural properties that distinguish it from the GSTs

of other families and it was identiﬁed as the prototype

of the beta class [14,26–36].

Three other orthologues of the beta class were also

functionally and structurally characterized, from

E. coli (EcGST) and from two soil bacteria, Ochrobac-

trum anthropi (OaGST) and Burkholderia xenovorans

(BxGST, also known as BphK), respectively

[20,21,24,37–47].

Theta class enzymes in bacteria are represented by

two dichloromethane (DCM) dehalogenases produced

by facultative methylotrophic bacteria [48–52]. They

exhibit high amino acid sequence similarity to eukar-

yotic theta class GSTs and also share some properties

of these enzymes such as their reactivity with DCM,

their lack of activity with CDNB and their inability to

bind to GSH afﬁnity matrices [50].

Tetrachlorohydroquinone (TCHQ) dehalogenase

was reported by Anandarajah et al. [53] to belong to

the zeta class on the basis of multiple sequence align-

ments. In particular, this enzyme is characterized, in

the GSH site, by the distinctive motif of zeta class

enzymes including two serine and a cysteine residues.

TCHQ dehalogenase is involved in two steps of the

biodegradation of pentachlorophenol and it also has

isomerase activity [53–56].

Recently a novel class of cGSTs, called chi class,

was proposed [10]. Two cyanobacterial cGSTs have

been puriﬁed and characterized from Thermosynechoc-

cus elongatus BP-1 (TeGST) and Synechoccus elongatus

PCC 6301 (SeGST). Although TeGST and SeGST

showed typical structural features of cGSTs, the results

presented here argued against their incorporation into

the beta class. In particular, these enzymes lack cyste-

ines completely indicating a possible different evolu-

tionary pathway for the cyanobacterial GSTs from the

beta class GSTs.

Like eukaryotic organisms, bacteria are character-

ized by multiple GST genes of widely divergent

sequences and unknown function [57]. In the E. coli

genome, in addition to the beta class GST [37] and to

Stringent starvation protein A, a RNA polymerase

with fold similarity to cGSTs [3], six GST homologues

have been identiﬁed [58]. The products of two of these

genes, YfcF and YfcG, exhibited GST- and GSH-

dependent peroxidase activities and were involved in

the defence against oxidative stress [59]. Pseudomonads

possess more than 10 GST genes [57]. In P. mirabilis

as well as in Proteus vulgaris three and four different

GSTs were identiﬁed, respectively [14,60]. Recently,

genomic analysis of the Gram-negative Shewanella

oneidensis revealed the presence of six GST-like genes

[61]. Two of these GST products showed high

sequence similarities to DCM dehalogenases.

Bacterial 2-hydroxychromene-2-carboxylic acid (HCCA)

isomerase is a GSH-dependent enzyme that is impli-

N. Allocati et al. Bacterial GSTs

FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS 59

cated in the naphtahalene degradation pathway

[5,62,63]. HCCA isomerase shares strong similarity

with kappa class enzymes, has a conserved serine resi-

due in the GSH site and displays catalytic activity

against CDNB [63]. Kappa class GSTs are soluble

dimeric proteins isolated and characterized from mam-

malian mitochondria [1,5]. In humans, hGSTK1 was

also located in the peroxisomal fraction suggesting a

new role for this class [1]. They are structurally distinct

from cGSTs, exhibit GSH activity towards haloarenes

and reduce cumene hydroperoxide and tert-butyl

hydroperoxide [1,5].

The MAPEG family are a wide and ubiquitous

group with diverse biological functions [6,64]. The

eukaryotic MAPEG family comprises at least six mem-

bers: 5-lipoxigenase activating protein (FLAP) and leu-

kotriene C

synthase (LTC

) involved in leukotriene

biosynthesis, MGST1, MGST2 and MGST3 with GST

and peroxidase activity and prostaglandin E synthase

(PGES) which catalyses the formation of prostaglan-

din E

from prostaglandin H

[6,64,65]. Recently,

crystallographic studies on different MAPEG members

clearly demonstrated that they are arranged into

trimers [66].

MAPEG members were also identiﬁed in several

bacteria such as E. coli,Vibrio cholerae and Synecho-

cystis sp. [6]. Some of them contained multiple MA-

PEG paralogues. They showed distant evolutionarily

relationships compared with eukaryotic mammalian

and non-mammalian forms. Bacterial MAPEG pro-

teins have been grouped in two distinct subfamilies,

one of which comprises E. coli and V. cholerae pro-

teins and the other the Synechocystis sp. protein which

more closely resembles enzymes belonging to the

MGST2/FLAP/LTC

subgroup [65]. Genes encoding

MAPEG proteins from E. coli and Synechocystis sp.

have been cloned and overexpressed. Membrane frac-

tions from cells overproducing E. coli MGST as well

as SynMGST showed GST activity with CDNB as sec-

ond substrate [65]. To date, no information about their

physiological role in bacteria is available.

A phylogenetic analysis using representative

prokaryotic and eukaryotic members of the GST

superfamily is shown in Fig. 1. The phylogenetic rela-

tionships among bacterial GSTs are consistent with

their known functional and structural features, show-

ing that they are distributed in different classes. The

beta class comprises the majority of bacterial GSTs

and enzymes belonging to the chi class are closely

related to this class. The two DCM dehalogenases are

clustered together with mammalian theta class

enzymes. TCHQ dehalogenases are closely related to

the zeta class. Agrobacterium tumefaciens GST shows

closer evolutionary relationship with mammalian alpha

class than with bacterial GSTs as reported previously

[67]. Sphingomonas paucimobilis LigF and LigG group

together and are distant from the Sph. paucimobilis

LigE enzyme which is more closely related to the zeta

class enzymes. Finally, Rhodococcus AD45 form a new

branch and it is not grouped with any other class.

Fig. 1. Evolutionary relationship between representative bacterial and eukaryotic GSTs. Multiple sequence alignment was produced by using

CLUSTAL W2 [143] and improved manually. The unrooted phylogenetic trees were constructed by the neighbour-joining method, based on the

distances derived from the Dayhoff matrix, with MEGA 4.0 software [144]. The robustness of the branches was assessed by the bootstrap

method with 1000 replications. Only nodes with a bootstrap value > 25% are reported. The scale bar represents a distance of 0.5 substitu-

tions per site. The sequences have the following accession numbers: epsilon class: Anopheles gambiae (XP_319972), Drosophila melanogas-

ter (CG5164); delta class: An. gambiae (Q93113), Anopheles dirus (AF273039); theta class: D. melanogaster (Q9VG96), An. gambiae

(Q94999), Bos taurus (Q2NL00), Homo sapiens (P30712); Methylobacterium sp. DM4 (P21161); Methylophilus sp. DM11 (P43387); phi class:

Arabidopsis thaliana (P42769), Zea mays GSTF3 (Q9ZP62), Z. mays GSTF1 (P12653), Triticum aestivum (P30111); Sph. paucimobilis LigF

(P30347); Sph. paucimobilis LigG (BAA77216); lambda class: A. thaliana (Q9LZ07), Z. mays (P49248), A. thaliana GSTL2 (Q9M2W2); omega

class: H. sapiens (P78417), Rattus norvegicus (Q9Z339), Caenorhabditis elegans (AAA27959); tau class: Aegilops tauschii (O04941), Z. mays

GSTU1 (Y12862), Z. mays GSTU2 (AJ010439); Agrobacterium tumefaciens (Q8UJG9); alpha class: H. sapiens GSTA1 (P08263), Mus muscu-

lus, GSTA2 (P10648); R. norvegicus (P04904), Gallus gallus GSTA1 (Q08392), G. gallus GSTA2 (Q08393), Bos taurus (Q5E9G0); sigma class:

H. sapiens (O60760), R. norvegicus (O35543), C. elegans (O16116), Manduca sexta (P46429), Ommastrephes sloanei (P46088); mu class:

H. sapiens (P09488), M. musculus (P10649), G. gallus (P20136), Dermatophagoides pteronyssinus (P46419); pi class: Onchocerca volvulus

(P46427), Bufo bufo (P81942), H. sapiens (P09211), R. norvegicus (P04906); Rhodococcus AD45 (AJ249207); chi class: T. elongatus

(NP_680998), S. elongatus (YP_171005), beta class: Haemophilus inﬂuenzae (P44521), Xylella fastidiosa (Q9PE18), Xanthomonas campestris

(P45875), O. anthropi (P81065), Magnetospirillum magnetotacticum (ZP_00054555), B. xenovorans LB400 (Q9RAFO), P. mirabilis (P15214),

E. coli (P39100); Sph. paucimobilis LigE (BAA02032); zeta class: H. sapiens (O43708), M. musculus (Q9WVL0), T. aestivum (O04437), A. tha-

liana (Q9ZVQ3); Sphingobium chlorophenolicum (Q03520); Sphingomonas spUG30 (AY057901); rho class: Pleuronectes platessa (X63761),

Pagrus major (AB158412), Micropterus salmoides (AY335905), Branchiostoma belcheri tsingtaunese (AY279519); kappa class: H. sapiens

(Q9Y2Q3), R. norvegicus (P24473), Xenopus tropicalis (AAH87819), G. gallus (XP_416525), Ps. putida (Q51948). MAPEG: H. sapiens

(P10620), B. taurus (Q64L89), D. melanogaster (AAN85305), Tetraodon nigroviridis (CAF97117), Synechocystis sp. (P73795), Acaryochl-

oris marina (YP_001518348), E. coli (P64515), V. cholerae (NP_232423).

Bacterial GSTs N. Allocati et al.

60 FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS

Because kappa class and MAPEG proteins are evolu-

tionarily distant from cGSTs two separate trees are

presented as insets in the Fig. 1. Bacterial MAPEG

proteins group with eukaryotic members, whereas

Pseudomonas putida HCCA isomerase groups with

mammalian kappa class members. Fosfomycin

resistance proteins show high divergence in primary

sequence and in structure, thus they were not

considered.

This classiﬁcation is in agreement with the evolu-

tionary model proposed by Frova [11]. According to

this model, the ancestor from which GSTs originated

N. Allocati et al. Bacterial GSTs

FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS 61

is thioredoxin. The ﬁrst phase of the evolution of

GSTs from thioredoxins saw the addition at the

C-terminus, or the insertion between the thioredoxin

fold, of an all-helical domain to produce two inter-

mediates: bacterial glutaredoxin 2 (GRX2), which

evolved into the cGSTs; and bacterial disulﬁde-bond-

forming oxidoreductase A (DsbA), which evolved

into the kappa GSTs. In a second phase, cGSTs fur-

ther diversiﬁed into different classes. Beta class

enzymes evolved after the dimerization of GRX2.

This model is supported by the ability of beta class

enzymes to act as thiol disulﬁde oxidoreductases as

well as GSH-conjugating enzymes and by the pres-

ence of a cysteine residue in the GSH site [14,68,69].

Finally, a shift from cysteine to serine chemistry

resulted in the theta class, followed by the zeta class

and then by all others classes. To date, there have

are no clear hypotheses about the evolution of

MAPEG family members [11].

Structures

Most of the bacterial GSTs identiﬁed to date belong

to the bacterial-speciﬁc beta class and since 1998 the

crystal structures of four representatives of this class

have been determined, i.e. P. mirabilis GST (PmGST)

[12], E. coli GST (EcGST) [37], B. xenovorans BphK

(BxGST) [47] and O. anthropi GST (OaGST) [41].

This, together with extensive site-directed mutagenesis

analysis, has allowed us to dissect in detail the

structural and catalytic properties of beta class

enzymes.

Beta class GSTs are homodimers with a chain length

of 201–203 residues. They display the canonical

GST fold with a thioredoxin-like N-terminal domain

followed by an all-helical C-terminal domain separated

by a short linker (Fig. 2A).

Comparative analysis of the crystal structures reveals

that the overall fold in beta class GSTs is remarkably

well conserved. Root mean square deviations among

equivalent Cas are generally < 1.5 A

˚when protein

monomers are superimposed, even though sequence

identities can be < 35–40%. When beta class monomers

are superimposed onto cGSTs belonging to other classes

rmsd values increase, ranging from 1.85 to 2.67 A

˚; these

values indicate that the canonical fold has been substan-

tially maintained from bacteria to mammals. Also the

monomers’ orientation in the different dimers is remark-

ably conserved (Fig. 2B). For example, when superim-

posing the OaGST dimer to the PmGST, EcGST and

BxGST dimers, rmsd values of 1.614, 1.418 and 1.54 A

˚,

respectively, are obtained [41]. These values are very

close to those obtained by superimposing the monomers

alone suggesting that strict conservation of the relative

orientation of the monomers in the dimer is required for

function.

Although the monomer fold in beta class GSTs

closely resembles that of other cGSTs, the dimer inter-

face is quite different. In fact, in contrast to other

GST classes, there is no open V-shaped interface,

although a close-packed arrangement is found, shaped

by the presence of stabilizing contacts at both the base

and top of the helical pairing that builds the interface

[12]. Furthermore, the interface in the majority of GST

classes is mainly hydrophobic in nature, whereas in

beta class GSTs it is markedly polar. As a conse-

quence, the well-known lock-and-key motif of alpha,

pi and mu class GSTs, comprising a protruding aro-

matic residue of one monomer that ﬁts into a hydro-

phobic pocket in the other, is absent in beta class

GSTs [12,41]. Conversely, other structural motifs origi-

nally identiﬁed in mammalian GSTs are conserved in

beta class GSTs indicating their ancient origin. Among

them, an important stabilizing role is played by the

so-called N-capping box and hydrophobic staple motifs

at the N-terminus of the sixth helix in the C-terminal

domain that contribute to the interaction with the N-

terminal thioredoxin-like domain [36]. An additional

structural motif has been identiﬁed in beta class GSTs

that appears to be restricted to this class. This motif is

formed by a network of hydrogen bonds, located in

the proximity of the G-site, which zippers the terminal

helix of the C-terminal domain to the starting helix of

the thioredoxin-like domain. Partial disruption of this

motif has been shown to have dramatic consequences

on the both the stability and the functionality of

OaGST [41].

GSH binds to beta class GSTs in an extended fash-

ion consistent with what is observed in other classes.

Several interactions, both polar and hydrophobic, con-

tribute to its stabilization, including a hydrogen bond

with an aspartate residue from the other monomer

[12,41].

All beta class GSTs are characterized by the pres-

ence of a conserved cysteine residue located close to

the GSH sulfydryl group. Notably a mixed disulﬁde

bond was found in the structure of PmGST, with the

two sulfurs located at a distance of 2.2 A

˚(Fig. 2C)

[12]. This oxidized state, however, was not found in

the structures of OaGST [41] and BxGST [47], where

the average distance between sulfur atoms is 3.3 A

˚,

consistent with GSH being in the thiolate form and

sharing a hydrogen with the cysteine sulfur of the pro-

tein. In PmGST, two other residues are at hydrogen

bond distance from the GSH sulfydryl group, a

histidine (His106) and a serine (Ser9) (Fig. 2C). The

Bacterial GSTs N. Allocati et al.

62 FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS

histidine is conserved in other beta class GSTs,

whereas Ser9 is not conserved in OaGST, BxGST and

EcGST [37,41,47]. Interestingly, mutagenesis data on

PmGST and EcGST indicated that none of the above-

mentioned residues (Ser9, Cys10 and His106) is strictly

required for the classical conjugating activity or is

solely responsible for the reduced pK

of GSH, sug-

gesting that, in contrast with mammalian GSTs, the

stabilization of the GSH thiolate is likely the result of

a network of interactions [31,33,70]. OaGST deviates

partly from this behaviour. In fact, in OaGST the

G-site histidine adopts a different orientation with

respect to the other enzymes and does not bind GSH

[41]. In OaGST, in contrast to what is observed with

PmGST and EcGST, mutation of the conserved cyste-

ine to alanine led to a marked decrease in speciﬁc

activity, due to a dramatic loss in afﬁnity for GSH

[42]. Interestingly, it was shown that, in this mutant,

GSH preferentially binds in a non-canonical position,

occupying the H-site [42]. Another beta class enzyme,

BxGST, was also shown to be able to bind a GSH in

the H-site [47]. Remarkably this enzyme was crystal-

lized in the presence of two GSH molecules, one with

the canonical G-site orientation and the other bound

at the H-site (Fig. 2D). In this structure the G- and

H-site GSH sulfurs are close (4.5 A

˚). These struc-

tural data support a mechanism in which once a disul-

ﬁde bond is formed between the G-site GSH and

Cys10 in the reduction of several substrates, a second

GSH enters the H-site and a disulﬁde bond exchange

takes place with the formation and release of an

oxidized glutathione (GSSG), thus restoring the

enzyme’s functionality [47].

Two bacterial proteins show signiﬁcant sequence

identity with mitochondrial kappa class GSTs. They

are DsbA and the HCCA isomerase. Kappa class

enzymes are peculiar in that, although they contain a

thioredoxin domain and an all-helical domain, their

topology differs from that of cytosolic enzymes

because the all-helical domain is inserted into the

thioredoxin fold. DsbA is a structural but not a func-

tional homologue of kappa class GSTs because its

active site does not bind GSH and it contains a

CXXC motif to perform redox catalysis [71]. HCCA

isomerase is the fourth enzyme in the naphthalene

catabolic pathway of Ps. putida. It catalyses the

conversion of HCCA, derived from cis-o-hydroxyben-

zeylidene pyruvic acid, to the more stable trans-o-

hydroxybenzeylidene pyruvic acid (Fig. 3I). Recently,

this enzyme has been subjected to extensive structural

and functional characterization that has shed light on

its peculiarities [63]. HCCA isomerase is a bona ﬁde

Fig. 2. Structural studies on beta class

GSTs. (A) Structure of OaGST shown with

the twofold symmetry axis perpendicular to

the page (pdb code: 2NTO). The N-terminal

thioredoxin domain and the C-terminal all

helical domain are highlighted with different

tonalities. GSH molecules are shown in

sticks. (B) Overlay of four different beta

class GSTs dimers shown as Catraces: Oa-

GST (red), BxGST (green, pdb code: 2GDR),

EcGST (magenta, pdb code: 1A0F) and

PmGST (cyan, pdb code: 1PMT). This repre-

sentation highlights the conservation of beta

class fold as well as of the monomers’ ori-

entation in the dimers. (C) GSH binding site

in PmGST. In this crystal structure, GSH

(green carbons) forms a mixed disulﬁde with

Cys10. His106 and Ser9 are also at hydro-

gen bond distance from the GSH sulfur. (D)

Structure of BxGST in complex with two

GSH molecules (green carbons), one at the

canonical G-site and the other bound at the

hydrophobic substrates binding site (H-site).

The two GSH sulfurs are at close distance

from each other. This ﬁgure was prepared

with PYMOL (DeLano Scientiﬁc).

N. Allocati et al. Bacterial GSTs

FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS 63

Bacterial GSTs N. Allocati et al.

64 FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS

kappa class GST: it conserves the same topology as

the mammalian enzyme, it is able to bind GSH,

which is activated through a conserved serine residue,

and it performs the classic conjugating reaction on

CDNB. Its structure superimposes well onto the

mammalian kappa class enzyme with only minor dif-

ferences and overall rmsd values among equivalent

Caof 2.2 A

˚. GSH is bound and stabilized in a

similar manner to mammalian kappa class GSTs but

more tightly, due to two additional interactions in

HCCA isomerase. A possible mechanism for the

HCCA isomerase reaction has been proposed in

which GSH performs a nucleophilic attack on the

HCCA ring and is then released thus behaving as a

cofactor (see later).

Structural genomics initiatives are increasing our

understanding of the protein-fold evolution and often

lead to the functional annotation of proteins whose

role was previously undetectable or merely hypotheti-

cal, based on sequence alignments alone. This is the

case of the Atu5508 gene product of Ag. tumefaciens

whose structure has recently been determined [67].

The protein has a dimeric organization, it binds GSH

and displays the canonical GST fold even though its

sequence identity is < 20% in pairwise comparisons

with any previously characterized cGSTs. The protein

is also a functional GST because it is able to conju-

gate GSH to p-benzyl chloride, a GST substrate.

Although coming from a bacterium, protein-fold anal-

ysis and structure superpositions suggest that this

protein is more closely related to mammalian GSTs

than to other bacterial GSTs. In particular, it lacks

the hallmarks of beta class GSTs, including absence

of the catalytic cysteine and histidine in the G-site

and the presence of a hydrophobic lock-and-key

motif at the dimer interface. For these reasons it has

been proposed that this protein is the prototype of a

new class of bacterial GSTs that includes several close

homologues found analysing a large set of environ-

mental sequences in the environmental sequencing

project [67].

Functions

Bacterial GSTs are specialized in several detoxiﬁcation

processes. They are able to detoxify a large number of

molecules via GSH conjugation (an example with the

classic substrate CDNB is shown in Fig. 3A). They

have an active role in protecting against oxidative

stress and are involved in the detoxiﬁcation of antimi-

crobial agents. Some are implicated in the basal

metabolism and supply bacterial cells with carbon

sources. Bacterial GSTs are also involved in the

degradation of several monocyclic aromatic com-

pounds such as toluene, xylenes, phenols and atrazine

[72]. They also take part in the degradation of poly-

cyclic aromatic hydrocarbons, a class of hazardous

chemicals to both environmental and human health

[21,73–78].

Oxidative and xenobiotic stress

Beta class cGSTs are involved in detoxication reactions

against toxic effects of several xenobiotics [39,40,

79–81].

In a modulation study it was reported that, in a

P. mirabilis strain, PmGST contributed to protect the

cells against oxidative stress induced by hydrogen per-

oxide [80]. Increases in the level of mRNA transcrip-

tion and in protein expression levels were observed

when the bacterial cells were exposed to hydrogen per-

oxide. This result was conﬁrmed by the analysis of a

gstB gene knock-out in the same P. mirabilis strain

that was found to be much more sensitive to hydrogen

peroxide than the wild-type strain [80]. Similar results

were also obtained for E. coli GST [59].

The modulation of OaGST in O. anthropi in the

presence of different xenobiotics was also investigated.

Unlike PmGST, hydrogen peroxide did not inﬂuence

the induction of the enzyme. Atrazine caused an

increase in the expression of OaGST at low, non-toxic

concentrations, suggesting its involvement in atrazine

metabolism [39]. Instead, phenolic compounds induced

a marked dose-dependent enhancement of the enzy-

matic cellular levels correlated to the toxicity of the

molecules indicating a role of OaGST in cellular

protection [39]. These data were also corroborated by

the preponderant presence of the enzyme in the peri-

plasmic space when bacteria were exposed to 4-chloro-

phenol [40].

However, although OaGST was found in the peri-

plasm, no signal sequence for export by the general

secretory (Sec) pathway was found [40]. Recently, an

alternative pathway has also been described, the twin-

Fig. 3. Reactions catalysed by bacterial GSTs. (A) Canonical conjugation of CDNB as second substrate; (B) dehalogenation of DCM to form-

aldehyde; (C) reductive dehalogenation of TCHQ to trichlorohydroquinone and of trichlorohydroquinone to 2,6-dichlorohydroquinone; (D) cis–

trans isomerization of maleylacetoacetate; (E) dehalogenation of 3-chloro-HOPDA; (F) hypothetical metabolic route for atrazine; (G) opening

of epoxide ring and reductive removal of GSH in isoprene metabolism; (H) b-aryl ether cleavage pathway; (I) step 4 of naphthalene catabolic

pathway; (J) opening of the epoxide ring of fosfomycin.

N. Allocati et al. Bacterial GSTs

FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS 65

arginine translocation (Tat) system, which allows the

transfer of folded proteins into periplasmic space [82].

Proteins transported via Tat system possess a distinc-

tive motif (SRRXFLK) at their N-terminus. This motif

was also not present in the OaGST sequence [83]. Sim-

ilarly, no Sec or Tat signals were found in PmGST,

EcGST, BxGST and Atu5508 [83,84]. Therefore,

the mechanism of OaGST transport (and possibly that

of other GSTs) into the periplasm is uncharacterized

at present. Moreover, a prokaryotic GSH transporter

in E. coli has recently been identiﬁed [85]. This

strongly suggests the presence of GSH in the peri-

plasm, as hypothesized previously [86], and underlines

that bacterial GSTs also display activity in this

compartment.

PmGST is able to catalyse a glutaredoxin-like reac-

tion using cysteine S-sulfate and hydroxyethyl disulﬁde

as substrates [68]. The G-site cysteine residue is essen-

tial for this redox activity [68] and the mixed disulﬁde

observed in the PmGST crystal structure has been

suggested as an intermediate in glutaredoxin-like

reactions [68].

Bacterial GSTs in dehalogenation

Microbial dehalogenases play a key role in the biodeg-

radation of several chlorinated xenobiotics, both ali-

phatic and aromatic [87,88]. Halogenated compounds

are widely used in industry and constitute an impor-

tant group of environmental pollutants.

Bacterial GSTs catalyse different reactions using

GSH as a cofactor, i.e. DCM dehalogenases catalyse

the hydrolytic dechlorination of dichloromethane,

whereas TCHQ dehalogenase catalyses a reductive

dehalogenation reaction.

DCM dehalogenases

DCM dehalogenase is a GSH-dependent enzyme syn-

thesized by a number of facultative methylotrophic

bacteria that are able to utilize DCM as a sole carbon

and energy source [48]. In the ﬁrst step of degradation,

the enzyme dechlorinates DCM to formaldeyde and

inorganic chloride (Fig. 3B). DCM, a signiﬁcant envi-

ronmental contaminant, is widely used as an industrial

solvent. The properties of two DCMDs are well docu-

mented in Methylobacterium dichloromethanicum DM4

and in Methylophilus leisingeri DM11 [48–52]. Both de-

halogenases are closely related to eukaryotic theta class

GSTs (Fig. 1). In particular, like theta class GSTs,

DCMDs share a conserved serine residue at the N-ter-

minal domain that is essential for catalysis [89]. Never-

theless, on the basis of other criteria such as N-

terminal amino acid sequences, kinetic and immuno-

logical properties, they have been further subdivided

into two classes. One class is formed by group A

enzymes, including the DCM dehalogenases of Methy-

lobacterium dichloromethanicum DM4. By contrast,

M. leisingeri DM11 is the prototype for group B

enzymes. The most signiﬁcant difference between the

two groups lies in their kinetic properties. Under con-

ditions of substrate saturation, DM11 is signiﬁcantly

faster in dechlorination than DM4.

To date, no structure is known for DCMDs. A 3D

homology model of DCMD from the M. leisingeri

DM11 strain has been presented based on alignments

with GST members from the theta and the alpha class

[90].

TCHQ dehalogenase

TCHQ dehalogenase catalyses the reduction of TCHQ

to trichlorohydroquinone and then to dichlorohydro-

quinone during the biodegradation of pentachlorophe-

nol, a xenobiotic compound present in the

environment, utilized primarily as fungicide for wood

preservation [91–94]. The catalytic mechanism of the

reaction was exhaustively studied by Copley et al.

[54,56,95]. TCHQ dehalogenase was puriﬁed by

Sphingobium chlorophenolicum, a soil bacterium that

can grow on pentachlorophenol as a sole carbon

source [92,96]. TCHQ dehalogenase utilizes two GSH

molecules to reductively dechlorinate TCHQ to trichlo-

rohydroquinone and then dichlorohydroquinone

(Fig. 3C). First, TCHQ dehalogenase catalyses the

nucleophilic attack of GSH on the substrate forming a

GSH conjugate, and then converts the conjugate to

reduced products. A conserved cysteine is required to

this second step of the reaction through the formation

of a mixed disulﬁde with GSH [54,97].

TCHQ dehalogenase also has maleylacetoacetate

isomerase activity, and the conserved cysteine is also

required for this activity [53]. Maleylacetoacetate

isomerases are enzymes that catalyse the GSH-depen-

dent cis–trans isomerization of maleylacetoacetate to

fumarylacetoacetate during the catabolism of tyrosine

and phenylalanine (Fig. 3D). Although the overall

sequence identity between TCHQ dehalogenase and

maleylacetoacetate isomerase is low, the active site is

highly conserved and in both cases contains a catalytic

cysteine [53].

Because maleylacetoacetate isomerases are part of

an ancient degradation pathway, whereas TCHQ

dehalogenase is a more specialized enzyme, it was

supposed that TCHQ dehalogenases evolved from

maleylacetoacetate isomerases [53].

Bacterial GSTs N. Allocati et al.

66 FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS

Biphenyl/polychlorinated

biphenyldehalogenation

A cGST characterized from B. xenovorans strain

LB400, a biphenyl/polychlorinated biphenyl-degrading

microorganism, known as BphK or BxGST, is one of

the most extensively investigated bacterial GSTs

[21,43–47]. This enzyme, which belongs to the beta

class, is encoded by a gene (bphK) located within the

bph locus, which speciﬁes the degradation pathway of

biphenyl and chlorobiphenyl compounds. B. xenovo-

rans LB400 is able to attack a broad spectrum of poly-

chlorinated biphenyl congeners and is able to grow on

biphenyl as sole carbon source [21,43]. Mutagenic

studies indicated that although the bphK gene is not

essential for utilization of this carbon source its expres-

sion gives advantage to the strain in the utilization of

biphenyls [43]. Subsequently, it has been established

that BphK catalyses efﬁciently the dehalogenation of

3-chloro-2-hydroxy-6-oxo-6-phenyl-2,4-dienoate com-

pounds that are produced by the co-metabolism of

polychlorinated biphenyls (Fig. 3E) [46]. The proposed

reaction mechanism is similar to that described for

TCHQ dehalogenase [56]. Although less efﬁcient,

BphK enzyme also has dechlorination activity against

4-chlorobenzoate [44]. A bphK gene is also present in

Sphingomonas yanoikuyae B1 and its expression allows

Sph. yanoikuyae B1 to grow faster on m-toluate

[98,99].

Atrazine metabolism

GSTs are able to detoxify several classes of herbicides

including triazines, a class of compounds to which

atrazine, one of the most widely used herbicides,

belongs [100,101]. GSTs are involved in the ﬁrst step

of atrazine biodegradation with the removal of the

chlorine atom produced by atrazine–GSH conjugation

[100,101]. Dechlorination is followed by the stepwise

removal of isopropylamine and ethylamine groups by

dealkylation [102]. In bacteria, atrazine can be degra-

dated either by a microbial consortium or by a single

microorganism [103,104]. For instance, in Pseudomonas

ADP, atrazine is metabolized to cyanuric acid by three

enzymatic steps. The ﬁrst step is performed by atrazine

chlorohydrolase followed by two dealkylations [104].

O. anthropi is a soil bacterium that is able to grow

on atrazine utilizing it as source of carbon [105] and

which expresses a functional beta class GST [24]. In a

modulation study of OaGST in the presence of several

xenobiotics, Favaloro and co-workers also showed

an increase in the levels of enzyme when atrazine

was added to the exponentially growing cells of

O. anthropi, suggesting an involvement of OaGST in

atrazine conjugation with GSH [39]. A proposed atra-

zine degradation pathway by bacterial GSTs is showed

in Fig. 3F.

Isoprene metabolism

Two GSTs were puriﬁed from the isoprene-utilizing

bacterium Rhodococcus sp. strain AD45 and their func-

tional properties were characterized [106–108]. Both

enzymes are involved in the metabolism of isoprene, an

atmospheric reactive hydrocarbon that plays a role in

ozone, organic peroxides and carbonic monoxide for-

mation, and their genes are localized in the isoprene

degradation gene cluster [106,108]. The ﬁrst GST,

encoded by the isoI gene, catalyses the GSH-dependent

ring opening of isoprene monoxide, the primary oxida-

tion product of isoprene (Fig. 3G). The GSH conjugate

1-hydroxy-2-glutathionyl-2-methyl-3-butene is then oxi-

dized in two consecutive steps to 2-glutathionyl-2-

methyl-3-butenoic acid by a dehydrogenase (IsoH). The

way in which isoprene degradation proceeds has not

been fully characterized. A convincing hypothesis is

that 2-glutathionyl-2-methyl-3-butenoic acid could be

converted to the corresponding CoA-thioester by a

racemase expressed by isoG gene, to allow the second

GST, which is encoded by the isoJ gene, to remove the

GSH molecule [108,109]. It is thought that IsoJ might

catalyse the reductive removal of GSH using a second

GSH molecule in a similar fashion as the TCHQ dehal-

ogenase (Fig. 3G) [54,91,108,109]. A relevant difference

from TCHQ dehalogenase is that the GSH conjugate is

formed several steps before its reduction, because the

removal of the GSH from 1-hydroxy-2-glutathionyl-2-

methyl-3-butene is energetically unfeasible [109]. More-

over, IsoI/GST is able to degrade halogen epoxides

such as 1,2-dichloroepoxyethanes and epichlorohydrin

[106,107]. In Rhodococcus AD45 a novel GST, namely

IsoILR1, was recently characterized with activity

towards cis-1,2-dichloroethylene epoxide and epoxypro-

pane [110].

Lignin degradation pathway

In Sph. paucimobilis SYK-6, the role of three tandemly

located genes, ligE,ligF and ligG, involved in the pro-

cess of lignin degradation, a fundamental step for the

earth’s carbon cycle, has been described [111–113].

Sph. paucimobilis is able to degrade a wide variety of

lignin compounds including b-aryl ether. The b-aryl

ether cleavage is a fundamental step in lignin degrada-

tion, because this intermolecular linkage is the most

abundant in lignin. LigE and LigF are enantioselective

N. Allocati et al. Bacterial GSTs

FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS 67

GSTs that cleave the b-aryl ether linkage with con-

sumption of GSH [113]. LigG is instead a GSH lyase

that catalyses the elimination of GSH from the conju-

gate produced by LigF (Fig. 3H) [112,113].

Naphthalene metabolism

HCCA isomerase is a GSH-dependent enzyme, struc-

turally related to mammalian kappa class enzymes,

involved in the naphthalene degradation pathway

[5,62,63]. HCCA isomerase is also involved in naph-

thalene sulfonates and anthracene catabolism

[114,115]. Naphthalene catabolism consists of an upper

pathway and a lower pathway. In the upper pathway,

naphthalene is metabolized to salycilate in six steps.

HCCA isomerase is the fourth enzyme of this pathway

and catalyses cis–trans isomerization between HCCA

and trans-o-hydroxybenzylidene pyruvic acid (Fig. 3I).

GSH is thought to be involved in this reaction by its

covalent addition of HCCA. The addition alters the

hybridization state at C7 promoting a rotation around

the C7–C8 double bond [63].

Interaction with antibiotics

Several studies on the interaction of PmGST with dif-

ferent classes of antibiotics have been performed and a

possible role for the enzyme in antibiotic-resistance

proposed [29,116].

First, it was observed that the efﬁciency of a number

of antimicrobial drugs decreased notably in the pres-

ence of the puriﬁed enzyme in the medium culture, as

shown by increased minimal inhibitory concentration

values [16,116]. By contrast, the presence of mamma-

lian GSTs had no effect on antibiotic efﬁciency [16].

Second, studies on the interaction of PmGST with sev-

eral antibiotics indicated that the enzyme sequesters

antimicrobial drugs with avidity [29]. The effect of 18

different antibiotics as inhibitors of PmGST activity

using CDNB and GSH as substrates, were also mea-

sured. Four of them, namely minocycline, tetracycline,

rifamycin and nitrofurantoin, were strong inhibitors

with IC

values between 49 and 140 lm. These drugs

produced a signiﬁcant decrease in the k

cat

values of the

bacterial enzyme for both substrates [29]. Furthermore,

PmGST displays a protective action against antibiotics

also in vivo [80,117]. An increase in PmGST levels was

observed when bacteria were grown in the presence of

drugs [80]. Moreover, viability tests showed that a gst

null-mutant P. mirabilis strain was more sensitive to

antibiotics than the wild-type bacterium [80]. Finally,

crystallographic data highlighted the presence of a

hydrophobic cavity large enough to bind antibacterial

molecules located at the dimer interface [12]. These

results were strengthened by the preponderant periplas-

mic location of the enzyme in the periplasmic space

[81].

Fosfomycin resistance proteins

Fosfomycin ([1R,2S]-[1,2-epoxypropyl]-phosphonic acid)

is a bactericidal broad-spectrum antibiotic effective

against both Gram-negative and Gram-positive bacte-

ria. Fosfomycin inhibits the enzyme MurA (UDP-

NAG enolpyruvyl transferase), which catalyses the

transfer of enolpyruvate from phosphoenolpyruvate to

uridine diphospho-N-acetylglucosamine, the ﬁrst com-

mitted step of bacterial cell-wall biosynthesis. Resis-

tance is mainly chromosomal but resistance genes have

also been found on transmissible plasmids [118]. Resis-

tance to fosfomycin can be achieved by several differ-

ent mechanisms, including decreased uptake of the

antibiotic, overexpression or mutation of MurA, and

enzymatic modiﬁcation of the antibiotic [118,119].

FosA, FosB and FosX represent three mechanisti-

cally distinct classes of enzymes that confer resistance

to fosfomycin by adding GSH, l-cysteine or a hydro-

xyl group, respectively, to the oxirane ring of the

antibiotic, and inactivating it.

FosA was originally identiﬁed in strains carrying

fosfomycin-resistance plasmids obtained from clinical

isolates [120–122]. Suarez and co-workers described a

new mechanism of antibiotic resistance due to the

enzymatic modiﬁcation of fosfomycin [121,122]. Inacti-

vation of fosfomycin occurred by the formation of an

adduct between its carbon 1 atom and the sulfydryl

group of the GSH cysteine resulting in the opening of

the epoxide ring of the antibiotic (Fig. 3J). The reac-

tion was catalysed by an enzyme that was referred to

as a GST. The enzyme was puriﬁed and characterized.

It did not bind to the GSH–agarose matrix and did

not catalyse the reaction between GSH and CDNB,

indicating that this protein had different properties

from the canonical GSTs. The enzyme is a homodimer

of 32 kDa and its activity is dependent on the addition

of the Mn

cation [122]. In subsequent studies, Arm-

strong and co-workers demonstrated that FosA is a

metalloglutathione transferase related to glyoxalase I

and extradiol dioxygenases, members of the vicinal

oxygen chelate superfamily [4,123]. In addition, they

showed that each subunit of the homodimer contains a

mononuclear Mn

centre that interacts strongly with

the antibiotic and also that the enzyme requires a

monovalent cation K

for optimal catalytic activity

[123]. FosA is also encoded in the bacterial genomes

and the 3D structure of a genomically encoded FosA

Bacterial GSTs N. Allocati et al.

68 FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS

from the pathogen Ps. aeruginosa was described [124].

The protein fold showed similarity to members of vici-

nal oxygen chelate superfamily and consists of paired

babbb motifs that form a cupped-shaped cavity for the

metal ion-binding site. The K

ion is accommodated

in a loop located 6.5 A

˚from the Mn

cation [124].

The structure of FosA in complex with antiobiotic was

also obtained. Plasmid-encoded and genomically

encoded enzymes are very similar in structure. The

crystal structure of FosA from transposon Tn2921

maintains the same basic molecular arrangement

observed in genomically encoded FosA [125]. Recently,

based on structural data, the residues involved in the

binding of both fosfomycin and GSH substrates have

been identiﬁed and characterized by mutagenesis

[126,127].

FosB was originally identiﬁed in fosfomycin-resis-

tant Staphylococcus strains and the genes were encoded

by plasmids [128–130]. A second type of FosB was

identiﬁed in the genome of a Bacillus subtilis strain

and subsequently characterized [131]. The protein is a

dimeric metallothiol transferase related to FosA.

Unlike FosA, FosB utilizes Mg

as metal cofactor.

In addition, FosB uses l-cysteine rather than GSH as

thiol donor and is less efﬁcient than FosA [131]. This

is in agreement with previous studies because the GSH

molecule was not detectable in most of the Gram-posi-

tive bacteria tested, including Bacillus subtilis [132].

FosX is a Mn(II)-dependent fosfomycin speciﬁc

epoxide hydrolase. It catalyses the addition of a water

molecule to fosfomycin, thus inactivating it. The

enzyme is genomically encoded, it is found in several

microorganisms and it has been well characterized

in Mesorhizobium loti and Lysteria monocytogenes

[133,134]. The two enzymes showed signiﬁcant func-

tional differences. Listeria FosX is a good catalyst and

is responsible for high resistance to fosfomycin [133].

By contrast, M. loti FosX produces modest resistance

to the antibiotic and, consistently, its kinetic costants

are lower than those of Listeria FosX. In addition,

M. loti FosX also catalyses the addition of GSH to

the antibiotic even if with low efﬁciency. It has been

suggested that M. loti FosX may be an intermediate in

the evolution of fosfomycin resistance proteins that

plays some yet to be identiﬁed role in the catabolism

of phosphonates. The structure of FosX was also

determined for both enzymes [133,134]. The FosX

structures are closely related and the overlay with

FosA from Ps. aeruginosa [124] shows a large degree

of similarity. Unlike FosA, the enzymes do not contain

ion-binding site near the active site. The

most interesting aspect of these structures is the obser-

vation that fosfomycin binds to FosX enzymes in a

different orientation from that observed in the FosA

enzyme.

A new mechanism of fosfomycin inactivation was

described by Garcia et al. [135] in a fosfomycin-resis-

tant strain of Pseudomonas syringae. The bacterium

yielded an enzyme, FosC, that inactivated the anti-

biotic using ATP to phosphorylate fosfomycin in the

presence of Mg

. This ﬁnding was corroborated by

sequence alignments highlighting a region of partial

homology between FosC and the Mg-ATP binding

domains of AMP–ATP phosphotransferases. To date,

no relationship between FosC and the other fosfo-

mycin resistance proteins is known.

Potential applications of bacterial

GSTs

As previously described, bacterial GSTs are involved

in several types of chemical transformations and may

represent a versatile tool with a variety of biotechno-

logical applications, for example, in the ﬁeld of bio-

remediation, an economical alternative to conventional

physicochemical methods to clean up environmentally

contaminated sites. The relative ease of genetic manip-

ulations in bacteria and their ability to grow rapidly

constitute a further advantage.

Several studies have been carried out exploiting the

potential of GSTs using both puriﬁed proteins and

microorganism engineering, some of which are summa-

rized here.

An example of protein engineering using the DNA-

shufﬂing technique was shown by Mannervik et al.

[136]. They hybridized six alpha class GSTs of differing

mammalian origin obtaining chimeric enzymes with

improved catalytic properties and altered substrate

selectivity towards several noxious iodoalkanes.

Another example is in the engineering of fusion pro-

teins with several distinct enzymatic activities. For

example, a trifunctional enzyme with superoxide

dismutase, glutathione peroxidase and glutathione

transferase activities was recently generated [137]. This

recombinant chimeric enzyme was shown to be effec-

tive in scavenging reactive oxygen species, thus show-

ing that this approach may have several applications

in medicine as well as in environmental ﬁeld.

Another potential application lies in the preparation

of biosensors. These are detection systems widely used

to check contaminated environments that combine a

biological component with a detector element. Biosen-

sors are competitive systems in comparison to conven-

tional methods being inexpensive, easy to use and

characterized by high sensitivity and selectivity. For

example, a mammalian GST was used to develop an

N. Allocati et al. Bacterial GSTs

FEBS Journal 276 (2009) 58–75 ª2008 The Authors Journal compilation ª2008 FEBS 69

optical biosensor for detection of captan in contami-

nated waters [138]. Captan is used to control a broad

spectrum of plant pathogenic microorganisms and it is

a strong inhibitor of GSTs [139].

A large number of bacterial species have developed

the ability to degrade xenobiotics previously considered

to be non-degradable [140]. Examples of microbial con-

sortia combining the ability of two or more bacterial

species to metabolize one or more noxious molecules

have been described [141]. An alternative route is to

engineer a single bacterial strain to carry a complete

metabolic pathway that efﬁciently eliminates environ-

mental toxic compounds [110]. For example, Wood

et al. [110] engineered in E. coli a metabolic pathway to

improve the degradation of chlorinated ethenes, which

constitute a large group of toxic environmental pollu-

tants [142]. The degradation of these molecules is limited

by the accumulation of reactive intermediates epoxides.

The authors constructed a recombinant E. coli strain in

which a toluene ortho-monooxygenase from Burkholde-

ria cepacia G4, obtained by DNA shufﬂing, and a GST

from Rhodococcus AD45 with activity towards cis-1,2,-

dichloroethylene epoxide and epoxypropane, were

co-expressed [110]. The ability of GST to transform

epoxides in E. coli strain increased the mineralization of

cis-1,2,-dichloroethylene.

The examples provided above highlight the potential

biotechnological applications of using engineered

proteins and bacterial strains. In this respect, bacterial

GSTs, which are characterized by high stability and by

a wide variety of catalysed reactions, undoubtedly,

constitute an effective resource for the future.

Acknowledgements

This work was supported in part by grants from

the Ministero dell’Istruzione, dell’Universita

`e della

Ricerca (MIUR) of Italy.

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Glutathione metabolism and regulation in cyanobacteria

Chapter

Jan 2024

Cyanobacteria are the ancient, monophyletic, and oxygen-producing photosynthetic group of bacteria that are widely distributed in all possible habitats. They are the first organisms to encounter oxidative stress in response to which they have evolved enzymatic and nonenzymatic strategies to combat stress. One of the crucial defenses relies on nonenzymatic antioxidants called glutathione. These are cellular thiols formed by cysteine with the help of ATP-requiring enzymes. Glutathione protects cyanobacteria against abiotic stresses like UV-B irradiation, ROS species, and high light intensity. It also aids in tolerance against cold, heat, metal stress, and nutrient deprivation as well as induces the production of various antioxidants. Glutathione poses various roles, majorly virulence and pathogenesis, biofilm formation and disruption, synthesis of DNA, and protection against methylglyoxal and copper homeostasis. Apart from the major emphasis on glutathione synthesis, metabolism, degradation, and roles, the present book chapter encompasses the source and effects of oxidative stress and its cross-talk with iron homeostasis.

Macroalgal microbiomes unveil a valuable genetic resource for halogen metabolism

Article

Full-text available

Mar 2024

Background Macroalgae, especially reds (Rhodophyta Division) and browns (Phaeophyta Division), are known for producing various halogenated compounds. Yet, the reasons underlying their production and the fate of these metabolites remain largely unknown. Some theories suggest their potential antimicrobial activity and involvement in interactions between macroalgae and prokaryotes. However, detailed investigations are currently missing on how the genetic information of prokaryotic communities associated with macroalgae may influence the fate of organohalogenated molecules. Results To address this challenge, we created a specialized dataset containing 161 enzymes, each with a complete enzyme commission number, known to be involved in halogen metabolism. This dataset served as a reference to annotate the corresponding genes encoded in both the metagenomic contigs and 98 metagenome-assembled genomes (MAGs) obtained from the microbiome of 2 red (Sphaerococcus coronopifolius and Asparagopsis taxiformis) and 1 brown (Halopteris scoparia) macroalgae. We detected many dehalogenation-related genes, particularly those with hydrolytic functions, suggesting their potential involvement in the degradation of a wide spectrum of halocarbons and haloaromatic molecules, including anthropogenic compounds. We uncovered an array of degradative gene functions within MAGs, spanning various bacterial orders such as Rhodobacterales, Rhizobiales, Caulobacterales, Geminicoccales, Sphingomonadales, Granulosicoccales, Microtrichales, and Pseudomonadales. Less abundant than degradative functions, we also uncovered genes associated with the biosynthesis of halogenated antimicrobial compounds and metabolites. Conclusion The functional data provided here contribute to understanding the still largely unexplored role of unknown prokaryotes. These findings support the hypothesis that macroalgae function as holobionts, where the metabolism of halogenated compounds might play a role in symbiogenesis and act as a possible defense mechanism against environmental chemical stressors. Furthermore, bacterial groups, previously never connected with organohalogen metabolism, e.g., Caulobacterales, Geminicoccales, Granulosicoccales, and Microtrichales, functionally characterized through MAGs reconstruction, revealed a biotechnologically relevant gene content, useful in synthetic biology, and bioprospecting applications. 7KFSwtia72NY8FA2HCcej3Video Abstract

Early transcriptional changes of heavy metal resistance and multiple efflux genes in Xanthomonas campestris pv. campestris under copper and heavy metal ion stress

Article

Full-text available

Mar 2024
BMC MICROBIOL

Background Copper-induced gene expression in Xanthomonas campestris pv. campestris (Xcc) is typically evaluated using targeted approaches involving qPCR. The global response to copper stress in Xcc and resistance to metal induced damage is not well understood. However, homologs of heavy metal efflux genes from the related Stenotrophomonas genus are found in Xanthomonas which suggests that metal related efflux may also be present. Methods and Results Gene expression in Xcc strain BrA1 exposed to 0.8 mM CuSO4.5H2O for 15 minutes was captured using RNA-seq analysis. Changes in expression was noted for genes related to general stress responses and oxidoreductases, biofilm formation, protein folding chaperones, heat-shock proteins, membrane lipid profile, multiple drug and efflux (MDR) transporters, and DNA repair were documented. At this timepoint only the cohL (copper homeostasis/tolerance) gene was upregulated as well as a chromosomal czcCBA efflux operon. An additional screen up to 4 hrs using qPCR was conducted using a wider range of heavy metals. Target genes included a cop-containing heavy metal resistance island and putative metal efflux genes. Several efflux pumps, including a copper resistance associated homolog from S. maltophilia, were upregulated under toxic copper stress. However, these pumps were also upregulated in response to other toxic heavy metals. Additionally, the temporal expression of the coh and cop operons was also observed, demonstrating co-expression of tolerance responses and later activation of part of the cop operon. Conclusions Overall, initial transcriptional responses focused on combating oxidative stress, mitigating protein damage and potentially increasing resistance to heavy metals and other biocides. A putative copper responsive efflux gene and others which might play a role in broader heavy metal resistance were also identified. Furthermore, the expression patterns of the cop operon in conjunction with other copper responsive genes allowed for a better understanding of the fate of copper ions in Xanthomonas. This work provides useful evidence for further evaluating MDR and other efflux pumps in metal-specific homeostasis and tolerance phenotypes in the Xanthomonas genus. Furthermore, non-canonical copper tolerance and resistance efflux pumps were potentially identified. These findings have implications for interpreting MIC differences among strains with homologous copLAB resistance genes, understanding survival under copper stress, and resistance in disease management.

Functionally coherent transcriptional responses of Jatropha curcas and Pseudomonas fragi for rhizosphere mediated degradation of pyrene

Article

Full-text available

Jan 2024

Pyrene is an extremely hazardous, carcinogenic polycyclic aromatic hydrocarbon (PAH). The plant–microbe interaction between Pseudomonas fragi DBC and Jatropha curcas was employed for biodegradation of pyrene and their transcriptional responses were compared. The genome of P. fragi DBC had genes for PAH degrading enzymes i.e. dioxygenases and dehydrogenases, along with root colonization (trpD, trpG, trpE and trpF), chemotaxis (flhF and flgD), stress adaptation (gshA, nuoHBEKNMG), and detoxification (algU and yfc). The transcriptional expression of catA and yfc that respectively code for catabolic enzyme (catechol-1, 2-dioxygnase) and glutathione-s-transferase for detoxification functions were quantitatively measured by qPCR. The catA was expressed in presence of artificial root exudate with or without pyrene, and glucose confirming the non-selective approach of bacteria, as desired. Pyrene induced 100-fold increase of yfc expression than catA, while there was no expression of yfc in absence of pyrene. The transcriptome of plant roots, in presence of pyrene, with or without P. fragi DBC inoculation was analysed. The P. fragi DBC could upregulate the genes for plant growth, induced the systemic acquired resistance and also ameliorated the stress response in Jatropha roots.

Architecture and potential roles of a delta-class glutathione S-transferase in protecting honey bee from agrochemicals

Article

Dec 2023
CHEMOSPHERE

Microcystin-LR Biodestruction by Autochthonous Microbiota of Different Water Bodies in the Northwest of Russia

Article

Nov 2023

The chromosome-scale reference genome of mirid bugs (Adelphocorissuturalis) genome provides insights into omnivory, insecticide resistance, and survival adaptation

Article

Full-text available

Sep 2023
BMC BIOL

Background Adelphocoris suturalis (Hemiptera: Miridae) is a notorious agricultural pest, which causes serious economic losses to a diverse range of agricultural crops around the world. The poor understanding of its genomic characteristics has seriously hindered the establishment of sustainable and environment-friendly agricultural pest management through biotechnology and biological insecticides. Results Here, we report a chromosome-level assembled genome of A. suturalis by integrating Illumina short reads, PacBio, 10x Chromium, and Hi-C mapping technologies. The resulting 1.29 Gb assembly contains twelve chromosomal pseudomolecules with an N50 of 1.4 and 120.6 Mb for the contigs and scaffolds, respectively, and carries 20,010 protein-coding genes. The considerable size of the A. suturalis genome is predominantly attributed to a high amount of retrotransposons, especially long interspersed nuclear elements (LINEs). Transcriptomic and phylogenetic analyses suggest that A. suturalis-specific candidate effectors, and expansion and expression of gene families associated with omnivory, insecticide resistance and reproductive characteristics, such as digestion, detoxification, chemosensory receptors and long-distance migration likely contribute to its strong environmental adaptability and ability to damage crops. Additionally, 19 highly credible effector candidates were identified and transiently overexpressed in Nicotiana benthamiana for functional assays and potential targeting for insect resistance genetic engineering. Conclusions The high-quality genome of A. suturalis provides an important genomic landscape for further investigations into the mechanisms of omnivory, insecticide resistance and survival adaptation, and for the development of integrated management strategies.

Contribution of the delta-class glutathione S-transferase to agrochemical adaptation in Apis mellifera

Preprint

Full-text available

Jul 2023

The European honey bee, Apis mellifera , serves as the principle managed pollinator species globally. In recent decades, honey bee populations have been facing serious health threats from combined biotic and abiotic stressors, including diseases, limited nutrition, and agrochemical exposure. Understanding the molecular mechanisms underlying xenobiotic adaptation of A. mellifera is critical, considering its extensive exposure to phytochemicals and agrochemicals present in flowers, propolis, hives, and the environment. In this study, we conducted a comprehensive structural and functional characterization of AmGSTD1, a delta class glutathione S-transferase (GST) enzyme, to unravel its roles in agrochemical detoxification and antioxidative stress responses. Significantly, we determined the 3D structure of a honey bee GST using protein crystallography for the first time, providing new insights into its molecular structure. Our investigations revealed that AmGSTD1 efficiently metabolizes model substrates, including 1-chloro-2,4-dinitrobenzene (CDNB), p-nitrophenyl acetate (PNA), phenylethyl isothiocyanate (PEITC), propyl isothiocyanate (PITC), and the oxidation byproduct 4-hydroxynonenal (4-HNE). Moreover, we discovered that AmGSTD1 exhibits binding affinity with the fluorophore 8-Anilinonaphthalene-1-sulfonic acid (ANS), which can be inhibited with various herbicides, fungicides, insecticides, and their metabolites. These findings highlight the potential contribution of AmGSTD1 in safeguarding honey bee health against various agrochemicals and their metabolites, while also mitigating oxidative stress resulting from exposure to these substances.

The plant stress hormone jasmonic acid evokes defensive responses in streptomycetes

Article

Full-text available

Oct 2023
APPL ENVIRON MICROB

Actinobacteria are prevalent in the rhizosphere and phyllosphere of diverse plant species where they help to enhance tolerance of plants against biotic and abiotic stresses. Here, we show that the plant hormones jasmonic acid (JA) and methyl jasmonate (MeJA) affect the growth, development, and specialized metabolism of Streptomyces . Exposure of Streptomyces coelicolor to JA or MeJA led to enhanced production of the polyketide antibiotic actinorhodin. JA also exhibited toxicity toward Streptomyces and Streptacidiphilus at higher concentrations, whereby streptomycetes were more tolerant to JA than members of the genus Streptacidiphilus . Tolerance to JA could be linked to its conjugation by the bacteria with glutamine. Additionally, JA conjugates with valine, tyrosine, phenylalanine, and leucine/isoleucine were identified. In contrast to JA, synthetic JA conjugates failed to activate antibiotic production and showed significantly reduced toxicity. Thus, our findings provide insights into a previously unknown defense mechanism deployed by Streptomycetaceae to a plant hormone. The underlying mechanism encompasses the attachment of amino acids to JA, which in turn safeguards the bacteria against the harmful impacts of the plant hormone. This study adds to the growing body of evidence that plant hormones can have a significant impact on members of the plant microbiome by affecting their growth, development, and secondary metabolism. IMPORTANCE Microorganisms that live on or inside plants can influence plant growth and health. Among the plant-associated bacteria, streptomycetes play an important role in defense against plant diseases, but the underlying mechanisms are not well understood. Here, we demonstrate that the plant hormones jasmonic acid (JA) and methyl jasmonate directly affect the life cycle of streptomycetes by modulating antibiotic synthesis and promoting faster development. Moreover, the plant hormones specifically stimulate the synthesis of the polyketide antibiotic actinorhodin in Streptomyces coelicolor . JA is then modified in the cell by amino acid conjugation, thereby quenching toxicity. Collectively, these results provide new insight into the impact of a key plant hormone on diverse phenotypic responses of streptomycetes.

Complete genome sequence of a plant growth-promoting endophytic bacterium V4 isolated from tea (Camellia sinensis) leaf

Article

Full-text available

Oct 2023

V4 is a Gram-negative, plant growth promoting endophytic bacterium that promotes the growth of tea plants. The appearance of V4 is rod shaped, with average dimensions of 1.34−1.5 × 0.32−0.39 μm and flagellum at both ends. The complete genome contains one circular chromosome and two plasmids. It is 4,697,109 bp in size, and contains 4,189 protein-coding genes, four gene islands and two prophages. Taxonomic classification suggested that V4 was a strain of Erwinia aphidicola. It was possible to find genes involved in plant growth promotion traits present in the genome of V4. Meanwhile, V4 was consistent with plant growth-promoting endophytic bacteria containing key synthetic genes associated with IAA synthesis, and P-solubilization, siderophores. V4 has siderophore biosynthesis genes compared with plant pathogenic bacteria showing stronger survival ability and the ability to interaction with the host plant. In addition, V4 endophytic bacteria possess a higher copy number of genes for flagellar assembly, bacterial chemotaxis and P-pilus assembly indicating stronger colonization and communication ability with host plants compared with five other bacteria in comparative genomic analysis. Analysis of the V4 endophytic bacterium complete genome sequence provides novel insights into the endophytic bacteria-host plant relationship, and suggests many candidate genes for post-genomic experiments.

Bacterial glutathione S-transferases: what are they good for?

Article

Full-text available

Mar 1997

Stéphane Vuilleumier

Structure, function and evolution of glutathione transferases: Implications for classification of non-mammalian members of an ancient enzyme superfamily

Article

Full-text available

Nov 2001

The glutathione transferases (GSTs; also known as glutathione S-transferases) are major phase II detoxification enzymes found mainly in the cytosol. In addition to their role in catalysing the conjugation of electrophilic substrates to glutathione (GSH), these enzymes also carry out a range of other functions. They have peroxidase and isomerase activities, they can inhibit the Jun N-terminal kinase (thus protecting cells against H(2)O(2)-induced cell death), and they are able to bind non-catalytically a wide range of endogenous and exogenous ligands. Cytosolic GSTs of mammals have been particularly well characterized, and were originally classified into Alpha, Mu, Pi and Theta classes on the basis of a combination of criteria such as substrate/inhibitor specificity, primary and tertiary structure similarities and immunological identity. Non-mammalian GSTs have been much less well characterized, but have provided a disproportionately large number of three-dimensional structures, thus extending our structure-function knowledge of the superfamily as a whole. Moreover, several novel classes identified in non-mammalian species have been subsequently identified in mammals, sometimes carrying out functions not previously associated with GSTs. These studies have revealed that the GSTs comprise a widespread and highly versatile superfamily which show similarities to non-GST stress-related proteins. Independent classification systems have arisen for groups of organisms such as plants and insects. This review surveys the classification of GSTs in non-mammalian sources, such as bacteria, fungi, plants, insects and helminths, and attempts to relate them to the more mainstream classification system for mammalian enzymes. The implications of this classification with regard to the evolution of GSTs are discussed.

The Atrazine Catabolism Genes atzABC Are Widespread and Highly Conserved

Article

Full-text available

Apr 1998

Pseudomonas strain ADP metabolizes the herbicide atrazine via three enzymatic steps, encoded by the genes atzABC, to yield cyanuric acid, a nitrogen source for many bacteria. Here, we show that five geographically distinct atrazine-degrading bacteria contain genes homologous to atzA, -B, and -C. The sequence identities of the atz genes from different atrazine-degrading bacteria were greater than 99% in all pairwise comparisons. This differs from bacterial genes involved in the catabolism of other chlorinated compounds, for which the average sequence identity in pairwise comparisons of the known members of a class ranged from 25 to 56%. Our results indicate that globally distributed atrazine-catabolic genes are highly conserved in diverse genera of bacteria.

Occurrence of glutathione in bacteria

Article

Mar 1978

Glutathione and soluble thiol content were examined in a broad spectrum of bacteria. Significant soluble thiol was present in all cases. The thiol compound was glutathione in most of the gram-negative bacteria but not in most of the gram-positive bacteria studied. Glutathione was absent in four anerobes and one microaerophile but was present in a blue-green bacterium. The glutathione content of Escherichia coli increased significantly during transition from exponential to stationary phase.

Glutathione S-Transferase from Escherichia coli

Article

Jan 1981

Takashi Shishido

Bioinformatic and enzymatic characterization of the MAPEG superfamily

Article

Apr 2005
FEBS J

The membrane associated proteins in eicosanoid and glutathione metabolism (MAPEG) superfamily includes structurally related membrane proteins with diverse functions of widespread origin. A total of 136 proteins belonging to the MAPEG superfamily were found in database and genome screenings. The members were found in prokaryotes and eukaryotes, but not in any archaeal organism. Multiple sequence alignments and calculations of evolutionary trees revealed a clear subdivision of the eukaryotic MAPEG members, corresponding to the six families of microsomal glutathione transferases (MGST) 1, 2 and 3, leukotriene C4 synthase (LTC4), 5-lipoxygenase activating protein (FLAP), and prostaglandin E synthase. Prokaryotes contain at least two distinct potential ancestral subfamilies, of which one is unique, whereas the other most closely resembles enzymes that belong to the MGST2/FLAP/LTC4 synthase families. The insect members are most similar to MGST1/prostaglandin E synthase. With the new data available, we observe that fish enzymes are present in all six families, showing an early origin for MAPEG family differentiation. Thus, the evolutionary origins and relationships of the MAPEG superfamily can be defined, including distinct sequence patterns characteristic for each of the subfamilies. We have further investigated and functionally characterized representative gene products from Escherichia coli, Synechocystis sp., Arabidopsis thaliana and Drosophila melanogaster, and the fish liver enzyme, purified from pike (Esox lucius). Protein overexpression and enzyme activity analysis demonstrated that all proteins catalyzed the conjugation of 1-chloro-2,4-dinitrobenzene with reduced glutathione. The E. coli protein displayed glutathione transferase activity of 0.11 micromol.min(-1).mg(-1) in the membrane fraction from bacteria overexpressing the protein. Partial purification of the Synechocystis sp. protein yielded an enzyme of the expected molecular mass and an N-terminal amino acid sequence that was at least 50% pure, with a specific activity towards 1-chloro-2,4-dinitrobenzene of 11 micromol.min(-1).mg(-1). Yeast microsomes expressing the Arabidopsis enzyme showed an activity of 0.02 micromol.min(-1).mg(-1), whereas the Drosophila enzyme expressed in E. coli was highly active at 3.6 micromol.min(-1).mg(-1). The purified pike enzyme is the most active MGST described so far with a specific activity of 285 micromol.min(-1).mg(-1). Drosophila and pike enzymes also displayed glutathione peroxidase activity towards cumene hydroperoxide (0.4 and 2.2 micromol.min(-1).mg(-1), respectively). Glutathione transferase activity can thus be regarded as a common denominator for a majority of MAPEG members throughout the kingdoms of life whereas glutathione peroxidase activity occurs in representatives from the MGST1, 2 and 3 and PGES subfamilies.

Effects of Cys10 mutation to Ala in glutathione transferase from Escherichia coli

Article

Oct 2000
J ORGANOMET CHEM

The residue Cys10 in E. coli glutathione transferase (GST) is apparently equivalent in primary structure to the catalytic Ser residue of the class theta GST and is located near the thiol group of GSH bound to the enzyme. The mutation of Cys10 to Ala, however, increased the specific activity toward 1-chloro-2,4-dinitrobenzene at pH 6.5. This mutation increased both the kcat and Km values for GSH and affected the pH-activity profile the enzyme. The side chain of Cys10 is thought to be important for construction of the GSH-binding site and partly for lowering the pKa of the GSH thiol, but not to be essential for the catalytic activity.

Identification of a Covalent Intermediate between Glutathione and Cysteine13 Formed during Catalysis by Tetrachlorohydroquinone Dehalogenase

Article

Nov 1997

Effect of Contaminant Concentration on Aerobic Microbial Mineralization of DCE and VC in Stream-Bed Sediments

Article

Jan 1998

Discharge of DCE and VC to an aerobic surface water system simultaneously represents a significant environmental concern and, potentially, a non-engineered opportunity for efficient contaminant bioremediation. The potential for bioremediation, however, depends on the ability of the stream-bed microbial community to efficiently and completely degrade DCE and VC over a range of contaminant concentrations. The purposes of the studies reported here were to assess the potential for aerobic DCE and VC mineralization by stream-bed microorganisms and to evaluate the effects of DCE and VC concentrations on the apparent rates of aerobic mineralization. Bed-sediment microorganisms indigenous to a creek, where DCE-contaminated groundwater continuously discharges, demonstrated rapid mineralization of DCE and VC under aerobic conditions. Over 8 days, the recovery of [1,2-14C]DCE radioactivity as 14CO2 ranged from 17% to 100%, and the recovery of [1,2-14C]VC radioactivity as 14CO2 ranged from 45% to 100%. Rates of DCE and VC mineralization increased significantly with increasing contaminant concentration, and the response of apparent mineralization rates to changes in DCE and VC concentrations was adequately described by Michaelis−Menten kinetics.

Common structural features of MAPEG - A widespread superfamily of membrane associated proteins with highly divergent functions in eicosanoid and glutathione metabolism

Article

Mar 1999
PROTEIN SCI

A novel superfamily designated MAPEG (Membrane Associated Proteins in Eicosanoid and Glutathione metabolism), including members of widespread origin with diversified biological functions is defined according to enzymatic activities, sequence motifs, and structural properties. Two of the members are crucial for leukotriene biosynthesis, and three are cytoprotective exhibiting glutathione S-transferase and peroxidase activities. Expression of the most recently recognized member is strongly induced by p53, and may therefore play a role in apoptosis or cancer development. In spite of the different biological functions, all six proteins demonstrate common structural characteristics typical of membrane proteins. In addition, homologues are identified in plants, fungi, and bacteria, demonstrating this superfamily to be generally occurring.

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