Out‐of‐plane deformations of the heme group in different ferrocytochrome c proteins probed by resonance Raman spectroscopy

Hagarman, Andrew; Wallace, Carmichael J. A.; Laberge, Monique; Schweitzer‐Stenner, Reinhard

doi:10.1002/jrs.2050

Cited by 14 publications

(11 citation statements)

References 51 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…58 These spectral changes are sensitive to the structure of the substrate and the mutation in the H76 residue, demonstrating that the structural properties of the heme are closely coupled to the substrate through the H76 residue.…”

Section: Resultsmentioning

confidence: 96%

Substrate−Protein Interaction in Human Tryptophan Dioxygenase: The Critical Role of H76

Batabyal

Yeh

2009

J. Am. Chem. Soc.

View full text Add to dashboard Cite

The initial and rate-limiting step of the kynurenine pathway in humans involves the oxidation of tryptophan to N-formyl kynurenine catalyzed by two hemeproteins, tryptophan 2,3-dioxygenase (hTDO) and indoleamine 2,3-dioxygenase (hIDO). In hTDO, the conserved H76 residue is believed to act as an active site base to deprotonate the indole NH group of Trp, the initial step of the Trp oxidation reaction. In hIDO, this histidine is replaced by a serine. To investigate the role of the H76, we have studied the H76S and H76A mutants of hTDO. Activity assays show that the mutations cause a decrease in kcat and an increase in KM for both mutants. The decrease in the kcat is accounted for by the replacement of the active site base catalyst, H76, with a weaker base, possibly a water, whereas the increase in KM is attributed to the loss of the specific interactions between the H76 and the substrate as well as the protein matrix. Resonance Raman studies with various Trp analogs indicate that the substrate is positioned in the active site by the ammonium, carboxylate, and indole groups, via intricate H-bonding and hydrophobic interactions. This scenario is consistent with the observation that L-Trp binding significantly perturbs the electronic properties of the O2-adduct of hTDO. The important structural and functional roles of H76 in hTDO is underscored by the observation that the electronic configuration of the active ternary complex, L-Trp-O2-hTDO, is sensitive to the H76 mutations.

show abstract

Section: Resultsmentioning

confidence: 96%

Substrate−Protein Interaction in Human Tryptophan Dioxygenase: The Critical Role of H76

Batabyal

Yeh

2009

J. Am. Chem. Soc.

View full text Add to dashboard Cite

show abstract

“…The much stronger intensity of the depolarized ν 15 band as compared to the polarized ν 7 band in WT is surprising. Recent studies conducted by Schweitzer-Stenner and coworkers suggest that the ν 15 band is a depolarized B 1g mode of a planar heme with D 4h symmetry; they showed that, if there is an in-plane distortion of the heme macrocycle, lowering the symmetry of the heme, the B 1g mode can be transformed into an A 1 mode, thereby gaining intensity in the resonance Raman spectrum [30]. Accordingly, we attribute the high relative intensity of the ν 15 band in the WT of PdC c O to an in-plane distortion of the heme a 3 , due to the interaction between the heme with its protein environment.…”

Section: Resultsmentioning

confidence: 99%

Modulation of the active site conformation by site-directed mutagenesis in cytochrome c oxidase from Paracoccus denitrificans

Ji¹,

Das²,

Puustinen³

et al. 2010

Journal of Inorganic Biochemistry

View full text Add to dashboard Cite

The structural and functional properties of active site mutants of cytochrome c oxidase from Paracoccus denitrificans (PdCcO) were investigated with resonance Raman spectroscopy. Based on the Fe-CO stretching modes and low frequency heme modes, two conformers (α- and β-forms) were identified that are in equilibrium in the enzyme. The α-conformer, which is the dominant species in the wild type enzyme, has a shorter heme a3 iron-CuB distance and a more distorted heme, as compared to the β-conformer, which has a more relaxed and open distal pocket. In general, the mutations caused a decrease in the population of the α-conformer, which is concomitant with a decreased in the catalytic activity, indicating that the α-conformer is the active form of the enzyme. The data suggest that the native structure of the enzyme is in a delicate balance of intramolecular interactions. We present a model in which the mutations destabilize the α-conformer, with respect to the β-conformer, and raise the activation barrier for the inter-conversion between the two conformers. The accessibility of the two conformers in the conformational space of CcO plausibly plays a critical role in coupling the redox reaction to proton translocation during the catalytic cycle of the enzyme.

show abstract

“…Notably, the type of OOP deformation is typically conserved within functionally-related classes of heme proteins 3. Ruffling is the principal OOP deformation in cytochrome c (cyt c ) electron transfer proteins,4–6 the enzyme-substrate complexes of heme oxygenases7 as well as the IsdG and IsdI heme-degrading enzymes,8 and the NO-carriers such as the nitrophorins and the heme nitric oxide/oxygen binding family of proteins 9–12. Saddling is prominent in peroxidases,13 while doming is typically observed in oxygen storage or transport proteins like myoglobin 14.…”

Section: Introductionmentioning

confidence: 99%

NMR and DFT Investigation of Heme Ruffling: Functional Implications for Cytochrome c

2010

View full text Add to dashboard Cite

Out-of-plane (OOP) deformations of the heme cofactor are found in numerous heme-containing proteins and the type of deformation tends to be conserved within functionally-related classes of heme proteins. We demonstrate correlations between the heme ruffling OOP deformation and the 13 C and 1 H nuclear magnetic resonance (NMR) hyperfine shifts of heme aided by density functional theory (DFT) calculations. The degree of ruffling in the heme cofactor of Hydrogenobacter thermophilus cytochrome c 552 has been modified by a single amino acid mutation in the second coordination sphere of the cofactor. The 13 C and 1 H resonances of the cofactor have been assigned using one-and two-dimensional NMR spectroscopy aided by selective 13 C-enrichment of the heme. DFT has been used to predict the NMR hyperfine shifts and electron paramagnetic resonance (EPR) g-tensor at several points along the ruffling deformation coordinate. The DFT-predicted NMR and EPR parameters agree with the experimental observations, confirming that an accurate theoretical model of the electronic structure and its response to ruffling has been established. As the degree of ruffling increases, the heme methyl 1 H resonances move upfield while the heme methyl and meso 13 C resonances move downfield. These changes are a consequence of altered overlap of the Fe 3d and porphyrin π orbitals, which destabilizes all three occupied Fe 3d-based molecular orbitals and decreases the positive and negative spin density on the β-pyrrole and meso carbons, respectively. Consequently, the heme ruffling deformation decreases the electronic coupling of the cofactor with external redox partners and lowers the reduction potential of heme.

show abstract

Out‐of‐plane deformations of the heme group in different ferrocytochrome c proteins probed by resonance Raman spectroscopy

Cited by 14 publications

References 51 publications

Substrate−Protein Interaction in Human Tryptophan Dioxygenase: The Critical Role of H76

Substrate−Protein Interaction in Human Tryptophan Dioxygenase: The Critical Role of H76

Modulation of the active site conformation by site-directed mutagenesis in cytochrome c oxidase from Paracoccus denitrificans

NMR and DFT Investigation of Heme Ruffling: Functional Implications for Cytochrome c

Contact Info

Product

Resources

About