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Modeling the backbone dynamics of reduced and oxidized solvated rat microsomal cytochrome b5.

ABSTRACT: In this article, a description of the statistics and dynamics of cytochrome b(5) in both reduced and oxidized forms is given. Results of molecular dynamics computer simulations in the explicit solvent have been combined with mode-coupling diffusion models including and neglecting the molecule-solvent correlations. R(1) and R(1 rho) nuclear magnetic relaxation parameters of (15)N in the protein backbone have been calculated and compared with experiments. Slight changes in charge density in the heme upon oxidation produces a cascade of changes in charge distributions from heme propionates up to charged residues approximately 1.5 nm from Fe. These changes in charge distributions modify the molecular surface and the water shell surrounding the protein. The statistical changes upon oxidation can be included in diffusive models that physically explain the upper and lower limits of R(1 rho) relaxation parameters at high off-resonance fields.

SUBMITTER: Giachetti A

PROVIDER: S-EPMC1304371 | biostudies-literature | 2004 Jul

REPOSITORIES: biostudies-literature

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Modeling the backbone dynamics of reduced and oxidized solvated rat microsomal cytochrome b5.

Giachetti Andrea A La Penna Giovanni La GL Perico Angelo A Banci Lucia L

Biophysical journal 20040701 1

In this article, a description of the statistics and dynamics of cytochrome b(5) in both reduced and oxidized forms is given. Results of molecular dynamics computer simulations in the explicit solvent have been combined with mode-coupling diffusion models including and neglecting the molecule-solvent correlations. R(1) and R(1 rho) nuclear magnetic relaxation parameters of (15)N in the protein backbone have been calculated and compared with experiments. Slight changes in charge density in the he ...[more]

PMID: 15240483

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Project description:Membrane cytochrome b5 reductase is a pleiotropic oxidoreductase that uses primarily soluble reduced nicotinamide adenine dinucleotide (NADH) as an electron donor to reduce multiple biological acceptors localized in cellular membranes. Some of the biological acceptors of the reductase and coupled redox proteins might eventually transfer electrons to oxygen to form reactive oxygen species. Additionally, an inefficient electron transfer to redox acceptors can lead to electron uncoupling and superoxide anion formation by the reductase. Many efforts have been made to characterize the involved catalytic domains in the electron transfer from the reduced flavoprotein to its electron acceptors, such as cytochrome b5, through a detailed description of the flavin and NADH-binding sites. This information might help to understand better the processes and modifications involved in reactive oxygen formation by the cytochrome b5 reductase. Nevertheless, more than half a century since this enzyme was first purified, the one-electron transfer process toward potential electron acceptors of the reductase is still only partially understood. New advances in computational analysis of protein structures allow predicting the intramolecular protein dynamics, identifying potential functional sites, or evaluating the effects of microenvironment changes in protein structure and dynamics. We applied this approach to characterize further the roles of amino acid domains within cytochrome b5 reductase structure, part of the catalytic domain, and several sensors and structural domains involved in the interactions with cytochrome b5 and other electron acceptors. The computational analysis results allowed us to rationalize some of the available spectroscopic data regarding ligand-induced conformational changes leading to an increase in the flavin adenine dinucleotide (FAD) solvent-exposed surface, which has been previously correlated with the formation of complexes with electron acceptors.

Project description:Formation of a transient [myoglobin (Mb), cytochrome b(5) (cyt b(5))] complex is required for the reductive repair of inactive ferri-Mb to its functional ferro-Mb state. The [Mb, cyt b(5)] complex exhibits dynamic docking (DD), with its cyt b(5) partner in rapid exchange at multiple sites on the Mb surface. A triple mutant (Mb(3M)) was designed as part of efforts to shift the electron-transfer process to the simple docking (SD) regime, in which reactive binding occurs at a restricted, reactive region on the Mb surface that dominates the docked ensemble. An electrostatically-guided brownian dynamics (BD) docking protocol was used to generate an initial ensemble of reactive configurations of the complex between unrelaxed partners. This ensemble samples a broad and diverse array of heme-heme distances and orientations. These configurations seeded all-atom constrained molecular dynamics simulations (MD) to generate relaxed complexes for the calculation of electron tunneling matrix elements (T(DA)) through tunneling-pathway analysis. This procedure for generating an ensemble of relaxed complexes combines the ability of BD calculations to sample the large variety of available conformations and interprotein distances, with the ability of MD to generate the atomic level information, especially regarding the structure of water molecules at the protein-protein interface, that defines electron-tunneling pathways. We used the calculated T(DA) values to compute ET rates for the [Mb(wt), cyt b(5)] complex and for the complex with a mutant that has a binding free energy strengthened by three D/E → K charge-reversal mutations, [Mb(3M), cyt b(5)]. The calculated rate constants are in agreement with the measured values, and the mutant complex ensemble has many more geometries with higher T(DA) values than does the wild-type Mb complex. Interestingly, water plays a double role in this electron-transfer system, lowering the tunneling barrier as well as inducing protein interface remodeling that screens the repulsion between the negatively-charged propionates of the two hemes.

Dataset Information

Modeling the backbone dynamics of reduced and oxidized solvated rat microsomal cytochrome b5.

Publications

Modeling the backbone dynamics of reduced and oxidized solvated rat microsomal cytochrome b5.

Similar Datasets

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