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A broken-symmetry density functional study of structures, energies, and protonation states along the catalytic O-O bond cleavage pathway in ba3 cytochrome c oxidase from Thermus thermophilus.

ABSTRACT: Broken-symmetry density functional calculations have been performed on the [Fea3, CuB] dinuclear center (DNC) of ba3 cytochrome c oxidase from Thermus thermophilus in the states of [Fea3(3+)-(HO2)(-)-CuB(2+), Tyr237(-)] and [Fea3(4+)[double bond, length as m-dash]O(2-), OH(-)-CuB(2+), Tyr237?], using both PW91-D3 and OLYP-D3 functionals. Tyr237 is a special tyrosine cross-linked to His233, a ligand of CuB. The calculations have shown that the DNC in these states strongly favors the protonation of His376, which is above propionate-A, but not of the carboxylate group of propionate-A. The energies of the structures obtained by constrained geometry optimizations along the O-O bond cleavage pathway between [Fea3(3+)-(O-OH)(-)-CuB(2+), Tyr237(-)] and [Fea3(4+)[double bond, length as m-dash]O(2-)HO(-)-CuB(2+), Tyr237?] have also been calculated. The transition of [Fea3(3+)-(O-OH)(-)-CuB(2+), Tyr237(-)] ? [Fea3(4+)[double bond, length as m-dash]O(2-)HO(-)-CuB(2+), Tyr237?] shows a very small barrier, which is less than 3.0/2.0 kcal mol(-1) in PW91-D3/OLYP-D3 calculations. The protonation state of His376 does not affect this O-O cleavage barrier. The rate limiting step of the transition from state A (in which O2 binds to Fea3(2+)) to state PM ([Fea3(4+)[double bond, length as m-dash]O(2-), OH(-)-CuB(2+), Tyr237?], where the O-O bond is cleaved) in the catalytic cycle is, therefore, the proton transfer originating from Tyr237 to O-O to form the hydroperoxo [Fea3(3+)-(O-OH)(-)-CuB(2+), Tyr237(-)] state. The importance of His376 in proton uptake and the function of propionate-A/neutral-Asp372 as a gate to prevent the proton from back-flowing to the DNC are also shown.

SUBMITTER: Han Du WG

PROVIDER: S-EPMC4972664 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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A broken-symmetry density functional study of structures, energies, and protonation states along the catalytic O-O bond cleavage pathway in ba3 cytochrome c oxidase from Thermus thermophilus.

Han Du Wen-Ge WG Götz Andreas W AW Yang Longhua L Walker Ross C RC Noodleman Louis L

Physical chemistry chemical physics : PCCP 20160420 31

Broken-symmetry density functional calculations have been performed on the [Fea3, CuB] dinuclear center (DNC) of ba3 cytochrome c oxidase from Thermus thermophilus in the states of [Fea3(3+)-(HO2)(-)-CuB(2+), Tyr237(-)] and [Fea3(4+)[double bond, length as m-dash]O(2-), OH(-)-CuB(2+), Tyr237˙], using both PW91-D3 and OLYP-D3 functionals. Tyr237 is a special tyrosine cross-linked to His233, a ligand of CuB. The calculations have shown that the DNC in these states strongly favors the protonation o ...[more]

PMID: 27094074

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Project description:The Fea3(3+)···CuB(2+) dinuclear center (DNC) structure of the as-isolated oxidized ba3 cytochrome c oxidase (CcO) from Thermus thermophilus (Tt) is still not fully understood. When the proteins are initially crystallized in the oxidized state, they typically become radiolyticly reduced through X-ray irradiation. Several X-ray crystal structures of reduced ba3 CcO from Tt are available. However, depending on whether the crystals were prepared in a lipidic cubic phase environment or in detergent micelles, and whether the CcO's were chemically or radiolyticly reduced, the X-ray diffraction analysis of the crystals showed different Fea3(2+)···CuB(+) DNC structures. On the other hand, Mössbauer spectroscopic experiments on reduced and oxidized ba3 CcOs from Tt (Zimmermann et al., Proc. Natl. Acad. Sci. USA 1988, 85, 5779-5783) revealed multiple (57)Fea3(2+) and (57)Fea3(3+) components. Moreover, one of the (57)Fea3(3+) components observed at 4.2 K transformed from a proposed "low-spin" state to a different high-spin species when the temperature was increased above 190 K, whereas the other high-spin (57)Fea3(3+) component remained unchanged. In the current Article, in order to understand the heterogeneities of the DNC in both Mössbauer spectra and X-ray crystal structures, the spin crossover of one of the (57)Fea3(3+) components, and how the coordination and spin states of the Fea3(3+/2+) and Cu(2+/1+) sites relate to the heterogeneity of the DNC structures, we have applied density functional OLYP calculations to the DNC clusters established based on the different X-ray crystal structures of ba3 CcO from Tt. As a result, specific oxidized and reduced DNC structures related to the observed Mössbauer spectra and to spectral changes with temperature have been proposed. Our calculations also show that, in certain intermediate states, the His233 and His283 ligand side chains may dissociate from the CuB(+) site, and they may become potential proton loading sites during the catalytic cycle.

Project description:The fundamental chemistry underpinning aerobic life on Earth involves reduction of dioxygen to water with concomitant proton translocation. This process is catalyzed by members of the heme-copper oxidase (HCO) superfamily. Despite the availability of crystal structures for all types of HCO, the mode of action for this enzyme is not understood at the atomic level, namely how vectorial H(+) and e(-) transport are coupled. Toward addressing this problem, we report wild type and A120F mutant structures of the ba(3)-type cytochrome c oxidase from Thermus thermophilus at 1.8 Å resolution. The enzyme has been crystallized from the lipidic cubic phase, which mimics the biological membrane environment. The structures reveal 20 ordered lipid molecules that occupy binding sites on the protein surface or mediate crystal packing interfaces. The interior of the protein encloses 53 water molecules, including 3 trapped in the designated K-path of proton transfer and 8 in a cluster seen also in A-type enzymes that likely functions in egress of product water and proton translocation. The hydrophobic O(2)-uptake channel, connecting the active site to the lipid bilayer, contains a single water molecule nearest the Cu(B) atom but otherwise exhibits no residual electron density. The active site contains strong electron density for a pair of bonded atoms bridging the heme Fe(a3) and Cu(B) atoms that is best modeled as peroxide. The structure of ba(3)-oxidase reveals new information about the positioning of the enzyme within the membrane and the nature of its interactions with lipid molecules. The atomic resolution details provide insight into the mechanisms of electron transfer, oxygen diffusion into the active site, reduction of oxygen to water, and pumping of protons across the membrane. The development of a robust system for production of ba(3)-oxidase crystals diffracting to high resolution, together with an established expression system for generating mutants, opens the door for systematic structure-function studies.

Dataset Information

A broken-symmetry density functional study of structures, energies, and protonation states along the catalytic O-O bond cleavage pathway in ba3 cytochrome c oxidase from Thermus thermophilus.

Publications

A broken-symmetry density functional study of structures, energies, and protonation states along the catalytic O-O bond cleavage pathway in ba3 cytochrome c oxidase from Thermus thermophilus.

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