Azide binding to yeast cytochrome c peroxidase and horse metmyoglobin: comparative thermodynamic investigation using isothermal titration calorimetry

Jacobson, Timothy; Williamson, Joy; Wasilewski, Anthony; Felesik, Janet; Vitello, Lidia B.; Erman, James E.

doi:10.1016/j.abb.2003.12.016

Cited by 12 publications

(10 citation statements)

References 43 publications

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“…These observations were further supported by the pK a values calculated for this residue before and after binding in each of these interactions. The pK a values calculated in the present study are in good accordance with values observed for histidines involved in catalysis (23)(24)(25)(26)(27) and macromolecular complex formation (28,29), which makes histidine the most likely candidate to be assigned for the protonation changes at the C3-compstatin interface. V4H9 has two histidines at positions 9 and 10, whereas V4W/H9A has only one histidine at position 10, and the other histidine at position 9 is replaced by an uncharged alanine residue.…”

Section: Discussionsupporting

confidence: 89%

Thermodynamic Studies on the Interaction of the Third Complement Component and Its Inhibitor, Compstatin

Katragadda

Morikis

Lambris

2004

Journal of Biological Chemistry

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Compstatin is a 13-residue cyclic peptide that inhibits complement activation by binding to complement component, C3. Although the activity of compstatin has been improved severalfold using combinatorial and rational design approaches, the molecular basis for its interaction with C3 is not yet fully understood. In the present study, isothermal titration calorimetry was employed to dissect the molecular forces that govern the interaction of compstatin with C3 using four different compstatin analogs. Our studies indicate that the C3-compstatin interaction is an enthalpy-driven process. Substitution of the valine and histidine residues at positions 4 and 9 with tryptophan and alanine, respectively, resulted in the increase of enthalpy of the interaction, thereby increasing the binding affinity for C3. The data also suggest that the interaction is mediated by water molecules. These interfacial water molecules could be the source for unfavorable entropy and large negative heat capacity changes observed in the interaction. Although part of the negative heat capacity changes could be accounted for by the water molecules, the rest might be resulting from the conformational changes in C3 and/or compstatin up on binding. Finally, we propose based on the pK a values determined from the protonation studies that histidine on compstatin participates in protonation changes and contributes to the specificity of the interaction between compstatin and C3. These protonation changes vary significantly between the binding of different compstatin analogs to C3.

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Section: Discussionsupporting

confidence: 89%

Thermodynamic Studies on the Interaction of the Third Complement Component and Its Inhibitor, Compstatin

Katragadda

Morikis

Lambris

2004

Journal of Biological Chemistry

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“…The fate of the released proton affects the pH dependence of binding. MetMb releases the acidic proton from ligands such as the imidazolium ion, HF, HCN, and HN 3 into solution [6-8] while the proton is retained within the peroxidase complexes, binding to the distal histidine [8,53]. For MIM binding to CcP, we suggest that there is little discrimination between the neutral base and the imidazolium cation in terms of entry into the distal heme pocket and binding to the heme iron, similar to the findings for metMb.…”

Section: Discussionmentioning

confidence: 99%

“…Heme proteins in their Fe(III) redox state bind ligands that are weakly basic such as azide, cyanide, fluoride and imidazole [1]. We have a long-standing interest in elucidating the differences in ligand binding to metmyoglobin (metMb) and cytochrome c peroxidase (CcP), representatives of two different heme protein classes [2-8]. The pH dependence of ligand binding to metMb and CcP are substantially different, reflecting differences in the role of acidic and basic groups within the two proteins in the binding process.…”

Section: Introductionmentioning

confidence: 99%

Binding of imidazole, 1-methylimidazole and 4-nitroimidazole to yeast cytochrome c peroxidase (CcP) and the distal histidine mutant, CcP(H52L)

Erman

Chinchilla

Studer

et al. 2015

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

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Imidazole, 1-methylimidazole and 4-nitroimidazole bind to yeast cytochrome c peroxidase (yCcP) with apparent equilibrium dissociation constants (KDnormalapp) of 3.3 ± 0.4, 0.85 ± 0.11, and ~0.2 M, respectively, at pH 7. This is the weakest imidazole binding to a heme protein reported to date and it is about 120 times weaker than imidazole binding to metmyoglobin. Spectroscopic changes associated with imidazole and 1-methylimidazole binding to yCcP suggest partial ionization of bound imidazole to imidazolate. The pKa for ionization of bound imidazole is estimated to be 7.4 ± 0.2, about 7 units lower than that of free imidazole and about 3 units lower than imidazole bound to metmyoglobin. Equilibrium binding of imidazole to CcP(H52L) is biphasic with low- and high-affinity phases having KDnormalapp values of 9.5 ± 4.5 and 0.13 ± 0.04 M, respectively. CcP(H52L) binding of 1-methylimidazole is monophasic with an affinity similar to those of yCcP and rCcP. Binding of 1-methylimidazole to rCcP is associated with two kinetic phases, the initial binding complete within 10 s, followed by a process that is consistent with 1-methylimidazole binding to a cavity created by movement of Trp-191 from the interior of the protein to the surface. Both the equilibrium binding and kinetics of 1-methylimidazole binding to yCcP are pH dependent. yCcP has a four-fold increase in 1-methylimidazole binding affinity on decreasing the pH from 7.5 to 4.0, an observation that is unique among the many studies on binding of imidazole and imidazole derivatives to heme proteins.

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“…Two are explored here. Figure 2G shows titrations of CCP proteins with bound azide or fluoride (Edwards and Poulos, 1990; Jacobson et al, 2004). Table 1 shows mid point potentials shifted ~−30 mV by azide and ~−83 mV by fluoride.…”

Section: Resultsmentioning

confidence: 99%

Impact of proximal and distal pocket site-directed mutations on the ferric/ferrous heme redox potential of yeast cytochrome c peroxidase

Jensen

Goodin

2011

Theor Chem Acc

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Cytochrome-c-peroxidase (CCP) contains a five-coordinate heme active site. The reduction potential for the ferric to ferrous couple in CCP is anomalously low and pH dependent (Eo = ~−180 mV vs. S.H.E. at pH 7). The contribution of the protein environment to the tuning of the redox potential of this couple is evaluated using site directed mutants of several amino acid residues in the environment of the heme. These include proximal pocket mutation to residues Asp-235, Trp-191, Phe-202 and His-175, distal pocket mutation to residues Trp-51, His-52, and Arg-48; and a heme edge mutation to Ala-147. Where unknown, the structural changes resulting from the amino acid substitution have been studied by X-ray crystallography. In most cases, ostensibly polar or charged residues are replaced by large hydrophobic groups or alternatively by Ala or Gly. These latter have been shown to generate large, solvent filled cavities. Reduction potentials are measured as a function of pH by spectroelectrochemistry. Starting with the X-ray derived structures of CCP and the mutants, or with predicted structures generated by Molecular Dynamics (MD), predictions of redox potential changes are modeled using the Protein Dipoles Langevin Dipoles (PDLD) method. These calculations serve to model an electrostatic assessment of the redox potential change with simplified assumptions about heme iron chemistry, with the balance of the experimentally observed shifts in redox potential being thence attributed to changes in the ligand set and heme coordination chemistry, and/or other changes in the structure not directly evident in the X-ray structures (e.g. ionization states, specific roles played by solvent species, or conformationally flexible portions of the protein). Agreement between theory and experiment is good for all mutant proteins with the exception of the mutation Arg 48 to Ala, and His 52 to Ala. In the former case, the influence of phosphate buffer is adduced to account for the discrepancy, and measurements made in a bis-tris propane/2-(N-morpholino)ethanesulfonic acid buffer system agree well with theory. For the latter case, an unknown structural element relevant to His-52, and/or solvent influence in the mutant akin to anion binding in the distal pocket (though lacking proof that it is) manifests in this mutant. The use of exogenous (sixth) ligands in dissecting the contributions to control of redox potential are also explored as a pathway for model building.

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Azide binding to yeast cytochrome c peroxidase and horse metmyoglobin: comparative thermodynamic investigation using isothermal titration calorimetry

Cited by 12 publications

References 43 publications

Thermodynamic Studies on the Interaction of the Third Complement Component and Its Inhibitor, Compstatin

Thermodynamic Studies on the Interaction of the Third Complement Component and Its Inhibitor, Compstatin

Binding of imidazole, 1-methylimidazole and 4-nitroimidazole to yeast cytochrome c peroxidase (CcP) and the distal histidine mutant, CcP(H52L)

Impact of proximal and distal pocket site-directed mutations on the ferric/ferrous heme redox potential of yeast cytochrome c peroxidase

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