Article

Crystallographic analysis of the interaction of nitric oxide with quaternary-T human hemoglobin.

Department of Biochemistry, College of Medicine, The University of Iowa, Iowa City, IA 52242, USA.
Biochemistry (Impact Factor: 3.38). 02/2004; 43(1):118-32. DOI:10.1021/bi030172j
Source: PubMed

ABSTRACT In addition to interacting with hemoglobin as a heme ligand to form nitrosylhemoglobin, NO can react with cysteine sulfhydryl groups to form S-nitrosocysteine or cysteine oxides such as cysteinesulfenic acid. Both modes of interaction are very sensitive to the quaternary structure of hemoglobin. To directly view the interaction of NO with quaternary-T deoxyhemoglobin, crystallographic studies were carried out on crystals of deoxyhemoglobin that were exposed to gaseous NO under a variety of conditions. Consistent with previous spectroscopic studies in solution, these crystallographic studies show that the binding of NO to the heme groups of crystalline wild-type deoxyhemoglobin ruptures the Fe-proximal histidine bonds of the alpha-subunits but not the beta-subunits. This finding supports Perutz's theory that ligand binding induces tension in the alpha Fe-proximal histidine bond. To test Perutz's theory, deoxy crystals of the mutant hemoglobin betaW37E were exposed to NO. This experiment was carried out because previous studies have shown that this mutation greatly reduces the quaternary constraints that oppose the ligand-induced movement of the alpha-heme Fe atom into the plane of the porphyrin ring. As hypothesized, the Fe-proximal histidine bonds in both the beta- and the alpha-subunits remain intact in crystalline betaW37E after exposure to NO. With regard to S-nitrosocysteine or cysteine oxide formation, no evidence for the reaction of NO with any cysteine residues was detected under anaerobic conditions. However, when deoxyhemoglobin crystals are first exposed to air and then to NO, the appearance of additional electron density indicates that Cys93(F9)beta has been modified, most likely to cysteinesulfenic acid. This modification of Cys93(F9)beta disrupts the intrasubunit salt bridge between His146(HC3)beta and Asp94(FG1)beta, a key feature of the quaternary-T hemoglobin structure. Also presented is a reanalysis of our previous crystallographic studies [Chan, N.-L., et al. (1998) Biochemistry 37, 16459-16464] of the interaction of NO with liganded hemoglobin in the quaternary-R2 structure. These studies showed additional electron density at Cys93(F9)beta that was consistent with an NO adduct. However, for reasons discussed in this paper, we now believe that this adduct may be the Hb-S-N.-O-H radical intermediate and not Hb-S-N=O as previously suggested.

0 0
 · 
0 Bookmarks
 · 
43 Views
  • [show abstract] [hide abstract]
    ABSTRACT: Methanosarcina acetivorans is a strictly anaerobic non-motile methane-producing Archaea expressing protoglobin (Pgb) which might either facilitate O(2) detoxification or act as a CO sensor/supplier in methanogenesis. Unusually, Methanosarcina acetivorans Pgb (MaPgb) binds preferentially O(2) rather than CO and displays anticooperativity in ligand binding. Here, kinetics and/or thermodynamics of ferric and ferrous MaPgb (MaPgb(III) and MaPgb(II), respectively) nitrosylation are reported. Data were obtained between pH 7.2 and 9.5, at 22.0 °C. Addition of NO to MaPgb(III) leads to the transient formation of MaPgb(III)-NO in equilibrium with MaPgb(II)-NO(+). In turn, MaPgb(II)-NO(+) is converted to MaPgb(II) by OH(-)-based catalysis. Then, MaPgb(II) binds NO very rapidly leading to MaPgb(II)-NO. The rate-limiting step for reductive nitrosylation of MaPgb(III) is represented by the OH(-)-mediated reduction of MaPgb(II)-NO(+) to MaPgb(II). Present results highlight the potential role of MaPgb in scavenging of reactive nitrogen and oxygen species.
    Biochemical and Biophysical Research Communications 12/2012; · 2.41 Impact Factor
  • Source
    [show abstract] [hide abstract]
    ABSTRACT: NO binding to the T-state of human hemoglobin (HbA) induces the cleavage of the proximal His 30 bonds to the heme iron in the a-chains, whereas it leaves the chain b-hemes hexacoordinated. 31 The structure of the nitrosylated T-state of the W37Eb mutant (W37E) shows that the Fe-His87a 32 bond remains intact. Exactly how mutation affects NO binding and why tension is apparent only 33 in HbA a-heme remains to be elucidated. By means of density functional theory electronic structure 34 calculations and classical molecular dynamics simulations we provide an explanation for the poorly 35 understood NO binding properties of HbA and its W37E mutant. The data suggest an interplay 36 between electronic effects, tertiary structure and hydration site modifications in determining the 37 tension in the NO-ligated T-state HbA a-chain.
    FEBS letters 06/2013; 587(15):2393-8. · 3.54 Impact Factor
  • Source
    [show abstract] [hide abstract]
    ABSTRACT: Despite their high physiological relevance, haemoglobin crystal structures with NO bound to haem constitute less than 1% of the total ligated haemoglobins (Hbs) deposited in the Protein Data Bank. The major difficulty in obtaining NO-ligated Hbs is most likely to be related to the oxidative denitrosylation caused by the high reactivity of the nitrosylated species with O(2). Here, using Raman-assisted X-ray crystallography, it is shown that under X-ray exposure (at four different radiation doses) crystals of nitrosylated haemoglobin from Trematomus bernacchii undergo a transition, mainly in the β chains, that generates a pentacoordinate species owing to photodissociation of the Fe-NO bond. These data provide a physical explanation for the low number of nitrosylated Hb structures available in the literature.
    Acta Crystallographica Section D Biological Crystallography 01/2013; 69(Pt 1):137-140. · 14.10 Impact Factor

Nei-Li Chan