David L Tierney

Miami University, Оксфорд, Ohio, United States

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Publications (58)318.48 Total impact

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    ABSTRACT: Cobalt is a metal contaminant at high temperature radioactive waste disposal sites. Past studies have largely focused on mesophilic microorganisms to remediate cobalt, despite the presence of thermophilic microorganisms at such sites. In this study, Methanothermobacter thermautotrophicus, a thermophilic methanogen, was used to reduce Co(III) in the form of [Co(III)-EDTA]−. Bioreduction experiments were conducted in a growth medium with H2/CO2 as a growth substrate at initial Co(III) concentrations of 1, 2, 4, 7, and 10 mM. At low Co(III) concentrations (< 4 mM), a complete reduction was observed within a week. Wet chemistry, X-ray absorption near-edge structure (XANES) and electron paramagnetic resonance (EPR) analyses were all consistent in revealing the reduction kinetics. However, at higher concentrations (7 and 10 mM) the reduction extents only reached 69.8% and 48.5%, respectively, likely due to the toxic effect of Co(III) to the methanogen cells as evidenced by a decrease in total cellular protein at these Co(III) concentrations. Methanogenesis was inhibited by Co(III) bioreduction, possibly due to impaired cell growth and electron diversion from CO2 to Co(III). Overall, our results demonstrated the ability of M. thermautotrophicus to reduce Co(III) to Co(II) and its potential application for remediating 60Co contaminant at high temperature subsurface radioactive waste disposal sites.
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    ABSTRACT: Matrix metalloproteinase-1 (MMP-1) plays crucial roles in diseases-related physiologies and pathological processes in the human body. We report here solution studies of MMP-1, including characterization of a series of mutants designed to bind metal in either the catalytic site or the structural site (but not both). CD and fluorescence spectroscopy of the mutants demonstrate the importance of the structural Zn(II) to maintaining both secondary and tertiary structure, while UV-vis, NMR, EPR and EXAFS show its presence influences the catalytic metal ion's coordination number. The mutants allow us to demonstrate convincingly the preparation of a mixed-metal analog, CoCZnS-MMP-1, with Zn(II) in the structural site and Co(II) in the catalytic site. Stopped-flow fluorescence of the native, ZnCZnS-MMP-1, and the mixed-metal CoCZnS-MMP-1 analog, show that the internal fluorescence of a nearby Trp residue modulates with catalysis and can be used to monitor reactivity under a number of conditions.
    Biochemistry 05/2015; 54(23). DOI:10.1021/acs.biochem.5b00379 · 3.02 Impact Factor
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    ABSTRACT: Despite significant progress on iron reduction by thermophilic microorganisms, studies on their ability to reduce toxic metals are still limited, despite their common co-existence in high temperature environments (up to 70 °C). In this study, Methanothermobacter thermautotrophicus, an obligate thermophilic methanogen, was used to reduce hexavalent chromium. Experiments were conducted in a growth medium with H2/CO2 as substrate with various Cr6+ concentrations (0.2, 0.4, 1, 3, and 5 mM) in the form of potassium dichromate (K2Cr2O7). Time-course measurements of aqueous Cr6+ concentrations using 1,5-diphenylcarbazide colorimetric method showed complete reduction of the 0.2 and 0.4 mM Cr6+ solutions by this methanogen. However, much lower reduction extents of 43.6%, 13.0%, and 3.7% were observed at higher Cr6+ concentrations of 1, 3 and 5 mM, respectively. These lower extents of bioreduction suggest a toxic effect of aqueous Cr6+ to cells at this concentration range. At these higher Cr6+ concentrations, methanogenesis was inhibited and cell growth was impaired as evidenced by decreased total cellular protein production and live/dead cell ratio. Likewise, Cr6+ bioreduction rates decreased with increased initial concentrations of Cr6+ from 13.3 to 1.9 μM h-1. X-ray absorption near-edge structure (XANES) spectroscopy revealed a progressive reduction of soluble Cr6+ to insoluble Cr3+ precipitates, which was confirmed as amorphous chromium hydroxide by selected area electron diffraction pattern. However, a small fraction of reduced Cr occurred as aqueous Cr3+. Scanning and transmission electron microscope observations of M. thermautotrophicus cells after Cr6+ exposure suggest both extra- and intracellular chromium reduction mechanisms. Results of this study demonstrate the ability of M. thermautotrophicus cells to reduce toxic Cr6+ to less toxic Cr3+ and its potential application in metal bioremediation, especially at high temperature subsurface radioactive waste disposal sites, where the temperature may reach ∼70 °C.
    Geochimica et Cosmochimica Acta 11/2014; in press. DOI:10.1016/j.gca.2014.10.012 · 4.33 Impact Factor
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    ABSTRACT: This study examines metal binding to metallo-β-lactamase VIM-2, demonstrating the first successful preparation of a Co(II)-substituted VIM-2 analog. Spectroscopic studies of the half- and fully-metal loaded enzymes show that both Zn(II) and Co(II) bind cooperatively, where the major species present, regardless of stoichiometry, are apo- and di-Zn (or di-Co) enzymes. We determined the di-Zn VIM-2 structure to a resolution of 1.55 Å, and this structure supports results from spectroscopic studies. Kinetics, both steady-state and pre-steady-state, show that VIM-2 utilizes a mechanism that proceeds through a very short-lived anionic intermediate when chromacef is used as the substrate. Comparison with other B1 enzymes shows that those that bind Zn(II) cooperatively are better poised to protonate the intermediate on its formation, compared to those that bind Zn(II) non-cooperatively, which uniformly build up substantial amounts of the intermediate.
    Biochemistry 10/2014; 53(46). DOI:10.1021/bi500916y · 3.02 Impact Factor
  • David L Tierney · Gerhard Schenk
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    ABSTRACT: In this mini-review, we briefly discuss the physical origin of x-ray absorption spectroscopy (XAS) before illustrating its application using dinuclear metallohydrolases as exemplary systems. The systems we have selected for illustrative purposes present a challenging problem for XAS, one that is ideal to demonstrate the potential of this methodology for structure/function studies of metalloenzymes in general. When the metal ion is redox active, XAS provides a sensitive measure of oxidation-state-dependent differences. When the metal ion is zinc, XAS is the only spectroscopic method that will provide easily accessible structural information in solution. In the case of heterodimetallic sites, XAS has the unique ability to interrogate each metal site independently in the same sample. One of the strongest advantages of XAS is its ability to examine metal ion site structures with crystallographic precision, without the need for a crystal. This is key for studying flexible metal ion sites, such as those described in the selected examples, because it allows one to monitor structural changes that occur during substrate turnover.
    Biophysical Journal 09/2014; 107(6):1263-1272. DOI:10.1016/j.bpj.2014.07.066 · 3.97 Impact Factor
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    ABSTRACT: Zinc finger proteins that bind Zn(II) using a Cys2His2 coordination motif within a ββα protein fold are the most abundant DNA binding transcription factor domains in eukaryotic systems. These classic zinc fingers are typically unfolded in the apo state and spontaneously fold into their functional ββα folds upon incorporation of Zn(II). These metal-induced protein folding events obscure the free energy cost of protein folding by coupling the protein folding and metal-ion binding thermodynamics. Herein, we determine the formation constant of a Cys2His2/ββα zinc finger domain, the C-terminal finger of the Wilms' tumor suppressor protein (WT1-4), for the purposes of determining its free energy cost of protein folding. Measurements of individual conditional dissociation constants, Kd values, at pH values from 5 to 9 were determined using fluorescence spectroscopy by direct or competition titration. Potentiometric titrations of apo-WT1-4 followed by NMR spectroscopy provided the intrinsic pKa values of the Cys2His2 residues, and corresponding potentiometric titrations of Zn(II)-WT1-4 followed by fluorescence spectroscopy yielded the effective pKa(eff) values of the Cys2His2 ligands bound to Zn(II). The Kd, pKa, and pKa(eff) values were combined in a minimal, complete equilibrium model to yield the pH-independent formation constant value for Zn(II)-WT1-4, Kf(ML) value of 7.5 × 10(12) M(-1), with a limiting Kd value of 133 fM. This shows that Zn(II) binding to the Cys2His2 site in WT1-4 provides at least -17.6 kcal/mol in driving force to fold the protein scaffold. A comparison of the conditional dissociation constants of Zn(II)-WT1-4 to those from the model peptide Zn(II)-GGG-Cys2His2 over the pH range 5.0 to 9.0 and a comparison of their pH-independent Kf(ML) values demonstrates that the free energy cost of protein folding in WT1-4 is less than +2.1 kcal/mol. These results validate our GGG model system for determining the cost of protein folding in natural zinc finger proteins and support the conclusion that the cost of protein folding in most zinc finger proteins is ≤+4.2 kcal/mol, a value that pales in comparison to the free energy contribution of Zn(II) binding, -17.6 kcal/mol.
    Inorganic Chemistry 06/2014; 53(12). DOI:10.1021/ic500862b · 4.76 Impact Factor
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    ABSTRACT: A mononuclear Cu(II) chlorodiketonate complex was prepared, characterized, and found to undergo oxidative aliphatic carbon-carbon bond cleavage within the diketonate unit upon exposure to O2 at ambient temperature. Mechanistic studies provide evidence for a dioxygenase-type carbon-carbon bond cleavage reaction pathway involving trione and hypochlorite intermediates. Significantly, the presence of a catalytic amount of chloride ion accelerates the oxygen activation step via the formation of a Cu-Cl species, which facilitates monodentate diketonate formation and lowers the barrier for O2 activation. The observed reactivity and chloride catalysis is relevant to Cu(II) halide catalyzed reactions in which diketonates are oxidatively cleaved using O2 as the terminal oxidant. The results of this study suggest that anion coordination can play a significant role in influencing copper-mediated oxygen activation in such systems.
    Journal of the American Chemical Society 05/2014; 136(22). DOI:10.1021/ja502577b · 12.11 Impact Factor
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    ABSTRACT: In an effort to characterize the roles of each metal ion in metallo-beta-lactamase NDM-1, heterodimetallic analogs (CoCo-, ZnCo-, and CoCd-) of the enzyme were generated and characterized. UV-visible, 1H NMR, EPR and EXAFS spectroscopies were used to confirm the fidelity of the metal substitutions, including the presence of a homogeneous, heterodimetallic cluster, with a single-atom bridge. This marks the first preparation of a metallo-beta-lactamase, selectively substituted with a paramagnetic metal ion, Co(II), in either the Zn1 (CoCd-NDM-1) or in the Zn2 site (ZnCo-NDM-1), as well as both (CoCo-NDM-1). We then used these metal-substituted forms of the enzyme to probe the reaction mechanism, using steady-state and stopped-flow kinetics, stopped-flow fluorescence and rapid-freeze-quench EPR. Both metal sites show significant effects on the kinetic constants, and both paramagnetic variants (CoCd- and ZnCo-NDM-1) showed significant structural changes on reaction with substrate. These changes are discussed in terms of a minimal kinetic mechanism that incorporates all of the data.
    Journal of the American Chemical Society 04/2014; 136(20). DOI:10.1021/ja410376s · 12.11 Impact Factor
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    ABSTRACT: The binding of three closely related chelators: 5-hydroxy-2-methyl-4H-pyran-4-thione (allothiomaltol, ATM), 3-hydroxy-2-methyl-4H-pyran-4-thione (thiomaltol, TM), and 3-hydroxy-4H-pyran-4-thione (thiopyromeconic acid, TPMA) to the active site of human carbonic anhydrase II (hCAII) has been investigated. Two of these ligands display a monodentate mode of coordination to the active site Zn2+ ion in hCAII that is not recapitulated in model complexes of the enzyme active site. This unprecedented binding mode in the hCAII-thiomaltol complex has been characterized by both X-ray crystallography and X-ray spectroscopy. In addition, the steric restrictions of the active site force the ligands into a 'flattened' mode of coordination compared with inorganic model complexes. This change in geometry has been shown by density functional computations to significantly decrease the strength of the metal-ligand binding. Collectively, these data demonstrate that the mode of binding by small metal-binding groups (MBGs) can be significantly influenced by the protein active site. Diminishing the strength of the metal-ligand bond results in unconventional modes of metal coordination not found in typical coordination compounds or even carefully engineered active site models, and understanding these effects is critical to the rational design of inhibitors that target clinically relevant metalloproteins.
    Journal of the American Chemical Society 03/2014; 136(14). DOI:10.1021/ja500616m · 12.11 Impact Factor
  • Aliyar Mousavi · Amy R. Marts · David L. Tierney · Stephen E. Cabaniss
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    ABSTRACT: The structure of Zn(II) complexes with dissolved organic matter (DOM) is an important consideration in developing molecular-level models of Zn(II) speciation, but recent reports favoring the tetrahedral geometry differ from earlier findings that geometry was largely octahedral. In general, the presence of thiolate ligands favors the tetrahedral geometry, while O and N ligands favor the octahedral geometry. This work presents extended X-ray absorption fine structure (EXAFS) and X-ray absorption near-edge structure (XANES) spectroscopic results, indicating an octahedral geometry over the pH range of 5–9 for a freshwater DOM isolate. Changes in XANES derivatives as a function of pH can be explained in terms of ligand protonation and/or changing ligand groups. Tetrahedral Zn(II)–DOM geometry may be restricted to binding environments containing thiol groups.
    Toxicological and Environmental Chemistry 01/2013; 95(1). DOI:10.1080/02772248.2012.734620 · 0.83 Impact Factor
  • David L Tierney
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    ABSTRACT: NMR paramagnetic relaxation enhancements (PREs) of a series of structurally-characterized, trigonal bistrispyrazolylborate (Tp) chelates of high-spin Co(II), spanning 100 - 850 MHz in field, are reported. Prior knowledge of the metal-nucleus distances allows numerical extraction of position-dependent electron spin relaxation rates (tc-1) from direct measurement of the individual PREs of the four symmetry distinct protons in Co(Tp)2, using available closed-form expressions. The data for this electronically complex system where spin-orbit coupling defines the ground state electronic structure are analyzed in terms of the Solomon-Bloembergen-Morgan (SBM) relations, as well as available zero-field splitting limit theories. A simple angular correction is shown to be sufficient to reconcile the individual tc(T) data for the four classes of protons. The data identify a previously unrecognized dynamic Jahn-Teller effect in these historically important complexes, with a barrier of ~ 230 cm-1, pointing to a level of dynamics in trispyrazolylborate chemistry that has not been described before, and further show that it is the Jahn-Teller that is responsible for the PREs in fluid solution. A field-dependent component is also identified for the two protons nearest g||, which is suggested to arise due to Zeeman mixing of excited state character into the ground level.
    The Journal of Physical Chemistry A 10/2012; 116(45). DOI:10.1021/jp309245e · 2.69 Impact Factor
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    ABSTRACT: Irradiation of 3-hydroxyflavonolato (3-Hfl) complexes of MnII, CoII, NiII and CuII (1–4) at 300 nm under aerobic conditions results in dioxygenase-type reactivity and the formation of the corresponding divalent metal O-benzoylsalicylato (O-bs) complexes 8–11 and CO. The latter were characterized by using multiple methods, including elemental analysis, X-ray crystallography, NMR and/or EPR spectroscopy, mass spectrometry and IR spectroscopy. Compounds 1–4 serve as catalysts for the photoinduced reactivity of 3-hydroxyflavonol (3-HflH) to produce O-benzoylsalicylic acid as the major product. Spectroscopic studies (UV/Vis and 1H NMR) show that each O-benzoylsalicylato complex 8–11 reacts with one equiv. of 3-hydroxyflavonol to regenerate 1–4 and enable turnover reactivity. Unlike what is observed for free 3-HflH, photoinduced reactions involving 1–4 and excess flavonol show only minor amounts of flavonol isomerization reactivity. These results indicate that the presence of a metal ion, whether under stoichiometric or catalytic conditions, facilitates the photoinduced degradation of 3-HflH to produce a carboxylic acid and CO as products.
    Berichte der deutschen chemischen Gesellschaft 10/2012; 2012(29). DOI:10.1002/ejic.201200212 · 2.94 Impact Factor
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    ABSTRACT: In an effort to biochemically characterize metallo-β-lactamase NDM-1, we cloned, overexpressed, purified, and characterized several maltose binding protein (MBP)-NDM-1 fusion proteins with different N-termini (full-length, Δ6, Δ21, and Δ36). All MBP-NDM-1 fusion proteins were soluble; however, only one, MBP-NDM-1Δ36, exhibited high activity and bound 2 equiv of Zn(II). Thrombin cleavage of this fusion protein resulted in the truncated NDM-1Δ36 variant, which exhibited a k(cat) of 16 s(-1) and a K(m) of 1.1 μM when using nitrocefin as a substrate, bound 2 equiv of Zn(II), and was monomeric in solution. Extended X-ray absorption fine structure studies of the NDM-1Δ36 variant indicate the average metal binding site for Zn(II) in this variant consists of four N/O donors (two of which are histidines) and 0.5 sulfur donor per zinc, with a Zn-Zn distance of 3.38 Å. This metal binding site is very similar to those of other metallo-β-lactamases that belong to the B1 subclass. Pre-steady-state kinetic studies using nitrocefin and chromacef and the NDM-1Δ36 variant indicate that the enzyme utilizes a kinetic mechanism similar to that used by metallo-β-lactamases L1 and CcrA, in which a reactive nitrogen anion is stabilized and its protonation is rate-limiting. While they are very different in terms of amino acid sequence, these studies demonstrate that NDM-1 is structurally and mechanistically very similar to metallo-β-lactamase CcrA.
    Biochemistry 04/2012; 51(18):3839-47. DOI:10.1021/bi300056y · 3.02 Impact Factor
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    ABSTRACT: To successfully establish an infection, Acinetobacter baumannii must overcome the iron starvation and oxidative stress imposed by the human host. Although previous studies have shown that ATCC 19606T cells acquire iron via the acinetobactin-mediated siderophore system, little is known about intracellular iron metabolism and its relation to oxidative stress in this pathogen. Screening of an insertion library resulted in the isolation of the ATCC 19606T derivative 1644, which was unable to grow in iron-chelated media. Rescue cloning and DNA sequencing showed that the insertion inactivated a gene coding for an NfuA Fe-S cluster protein ortholog, without any effect on the expression of the acinetobactin system. The nfuA mutant was also more sensitive to hydrogen peroxide and cumene hydroperoxide than the parental strain. The iron chelation- and oxidative-stress-deficient responses of this mutant were corrected when complemented with either the ATCC 19606T parental allele or the Escherichia coli MG1655 nfuA ortholog. Furthermore, electron paramagnetic resonance (EPR) and inductively coupled plasma-atomic emission spectroscopy (ICP-AES) analyses showed that the ATCC 19606T NfuA ortholog has iron-binding properties compatible with the formation of [Fe-S] cluster protein. Ex vivo and in vivo assays using human epithelial cells and Galleria mellonella, respectively, showed that NfuA is critical for bacterial growth independent of their capacity to acquire iron or the presence of excess of free iron. Taken together, these observations indicate that the A. baumannii NfuA ortholog plays a role in intracellular iron utilization and protection from oxidative-stress responses that this pathogen could encounter during the infection of the human host.
    Journal of bacteriology 03/2012; 194(11):2884-93. DOI:10.1128/JB.00213-12 · 2.81 Impact Factor
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    ABSTRACT: The tris(3-phenyl-5-methyl-1,2,4-triazolyl)borate (Ttz(Ph,Me)) ligand provides intermediate steric bulk and forms predominantly bis(ligand) complexes of the form M(Ttz(Ph,Me))(2) with first row divalent transition metals (1(M), M = Zn, Cu, Ni, Co, Fe, Mn). Due to ligand field effects that are greatest with Ni and Cu, ligand rearrangement is favored with these metals and Cu(Ttz(Ph,Me)*)(2) (1(Cu)*) and (Ttz(Ph,Me)*)Ni(Ttz(Ph,Me)) (1(Ni)*) were isolated by selective recrystallization and fully characterized (* indicates a rearranged Ttz ligand with Ph and Me in swapped positions in one triazole ring). For comparison with Co(Ttz(Ph,Me))(2), the less bulky analogs (Ttz(H,H))(2)Co (4) and (Ttz(Me,Me))(2)Co (5) were studied by NMR and EPR spectroscopy, and 5 was crystallographically characterized. These complexes allow for a study of how slight changes in structure and electron donor properties (for Ni and Cu), as well as dramatic changes in steric bulk (for Co), influence the physical properties; specifically there are significant changes in the UV-Vis, EPR and NMR spectra. Bis(ligand) complexes predominate with all metals, but (Ttz(Ph,Me))Ni(OH(2))Cl (2) and (Ttz(Ph,Me))ZnBr (3) were also isolated and these show that Ttz(Ph,Me) is coordinatively flexible.
    Dalton Transactions 03/2012; 41(9):2774-87. DOI:10.1039/c2dt12029a · 4.20 Impact Factor
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    ABSTRACT: Cobalt and zinc binding by the subclass B1 metallo-β-lactamase BcII from Bacillus cereus is examined by X-ray absorption spectroscopy, at various levels of metal loading. The data show that a significant amount of the dinuclear enzyme is formed, even at substoichiometric levels of metal loading, whether the added metal is Zn(II) or Co(II). Increasing metal addition, from 0.5 to 1.0 to 2.0eq/mol of enzyme, are shown to result in a more ordered active site. While Zn(II) appears to show no preference for the Zn(1) (3H) or Zn(2) (DCH) sites, the extended X-ray absorption fine structure (EXAFS) suggests that Co(II) shows a slight preference for the DCH site at low levels of added Co(II). The results are discussed in the context of similar metal-binding studies of other B1 metallo-β-lactamases.
    Journal of inorganic biochemistry 01/2012; 111:182-6. DOI:10.1016/j.jinorgbio.2011.12.013 · 3.44 Impact Factor
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    ABSTRACT: RT-PCR and DNA microarrays were used to probe for Zn(II)-responsive genes in E. coli cells that were made Zn(II) deficient. Microarray data revealed 114 genes were significantly up-regulated and 146 genes were significantly down-regulated in Zn(II) deficient conditions. The three most up-regulated genes were (1) znuA, which encodes for a periplasmic protein known to be involved with Zn(II) import, (2) yodA, which encodes for a periplasmic protein with unknown function, and (3) ykgM, which encodes for a ribosomal protein that is thought to be a paralog of ribosomal protein L31. YodA was over-expressed and purified as a maltose binding protein (MBP) fusion protein and shown to tightly bind 4 equivalents of Zn(II). Metal analyses showed that MBP-YkgM does not bind Zn(II). On the other hand, MBP-L31 tightly binds 1 equivalent of Zn(II). EXAFS studies on MBP-L31 suggest a ligand field of 1 histidine, 1 cysteine, and 2 additional N/O scatterers. Site-directed mutagenesis studies suggest that Cys16 coordinates Zn(II) in MBP-L31 and that the other three cysteines do not bind metal. These results are discussed in light of Zn(II) starvation model that has been postulated for B. subtilis.
    Journal of inorganic biochemistry 12/2011; 111:164-72. DOI:10.1016/j.jinorgbio.2011.11.022 · 3.44 Impact Factor
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    M Patrick Hensley · David L Tierney · Michael W Crowder
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    ABSTRACT: Escherichia coli 70S ribosomes tightly bind 8 equiv of Zn(II), and EXAFS spectra indicate that Zn(II) may be protein-bound. Ribosomes were incubated with EDTA and Zn(II), and after dialysis, the resulting ribosomes bound 5 and 11 equiv of Zn(II), respectively. EXAFS studies show that the additional Zn(II) in the zinc-supplemented ribosomes binds in part to the phosphate backbone of the ribosome. Lastly, in vitro translation studies demonstrate that EDTA-treated ribosomes do not synthesize an active Zn(II)-bound metalloenzyme, while the as-isolated ribosomes do. These studies demonstrate that the majority of intracellular Zn(II) resides in the ribosome.
    Biochemistry 11/2011; 50(46):9937-9. DOI:10.1021/bi200619w · 3.02 Impact Factor
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    ABSTRACT: In an effort to probe for metal binding to metallo-β-lactamase (MβL) IMP-1, the enzyme was overexpressed, purified, and characterized. The resulting enzyme was shown to bind 2 equiv of Zn(II), exhibit significant catalytic activity, and yield EXAFS results similar to crystallographic data previously reported. Rapid kinetic studies showed that IMP-1 does not stabilize a nitrocefin-derived reaction intermediate; rather, the enzyme follows a simple Michaelis mechanism to hydrolyze nitrocefin. Metal-substituted and metal-reconstituted analogues of IMP-1 were prepared by directly adding metal ion stocks to metal-free enzyme, which was generated by dialysis versus EDTA. UV-vis studies on IMP-1 containing 1 equiv of Co(II) showed a strong ligand-to-metal charge transition at 340 nm, and the intensity of this feature increased when the second equivalent of Co(II) was added to the enzyme. EXAFS fits on IMP-1 containing 1 equiv of Co(II) strongly suggest the presence of a metal-metal interaction, and EPR spectra of the IMP-1 containing 1 and 2 equiv of Co(II) are very similar. Taken together, steady-state kinetic and spectroscopic studies suggest that metal binding to metal-free IMP-1 follows a positive-cooperative mode.
    Biochemistry 09/2011; 50(42):9125-34. DOI:10.1021/bi200839h · 3.02 Impact Factor
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    ABSTRACT: The EPR spectra of high-spin Co(II) systems typically consist of poorly resolved resonances, broadened by a combination of large g-anisotropy, sizable spin–orbit couplings and admixture of excited state character into the magnetic ground state. In this report, we present dual-mode EPR studies of a set of 4-, 5- and 6-coordinate Co(II) complexes, as well as cobalt-substituted carbonic anhydrase, with and without a bidentate chelator. The suite of samples shows surprisingly large signals in parallel mode that cannot be accounted for by imperfections in the isolation of the parallel and perpendicular polarizations. To our knowledge, this is the first report of parallel polarization EPR from a Kramers system. We suggest that the parallel mode signals arise from admixture of excited state character into the ground state, and that the methods employed here may provide a means of quantitatively assessing the extent of mixing.
    Applied Magnetic Resonance 08/2011; 40(4):501-511. DOI:10.1007/s00723-011-0225-5 · 1.17 Impact Factor

Publication Stats

1k Citations
318.48 Total Impact Points


  • 2005–2015
    • Miami University
      • Department of Chemistry and Biochemistry
      Оксфорд, Ohio, United States
  • 2007–2012
    • University of New Mexico
      • Department of Chemistry and Chemical Biology
      Albuquerque, New Mexico, United States
    • Columbia University
      • Department of Chemistry
      New York City, NY, United States
  • 2005–2006
    • University of Texas at Austin
      • Division of Medicinal Chemistry
      Austin, Texas, United States
  • 1998–2005
    • Northwestern University
      • • Department of Cell and Molecular Biology
      • • Department of Chemistry
      Evanston, Illinois, United States
  • 2002
    • New University of Lisbon
      • Faculty of Sciences and Technology
      Caparica, Distrito de Setubal, Portugal