High-resolution crystal structure of cytochrome P450cam

Poulos, T.L.; Finzel, B.C.; Howard, Andrew

doi:10.1016/0022-2836(87)90190-2

Cited by 1,363 publications

(1,165 citation statements)

References 36 publications

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“…The secondary structure code as determined by the DSSP program (Kabsch & Sander, 1983): T, reverse turn; G, 310-helix; S, bend; E, P-strand; B, residue hydrogen bonding to a P-strand; -, no secondary structure. References: IGDl (Skarzynski et al, 1987), IGOX (Lindqvist, 1989), IIlB (Finzel et al, 1984), IPCY (Guss & Freeman, 1983), ZAPR (Suguna et al, 1987), ZCAB (Kannan et al, 1984), 2CPP (Poulos et al, 1987). 20VO (Bode et al, 1985), 2PRK (Betzel et al, 1988), 3DFR (Bolin et al, 1982).…”

Section: Cluster 131mentioning

confidence: 99%

Differences in the amino acid distributions of 3₁₀‐helices and α‐helices

1992

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Local determinants of 3,0-helix stabilization have been ascertained from the analysis of the crystal structure data base. We have clustered all 5-length substructures from 5 1 nonhomologous proteins into classes based on the conformational similarity of their backbone dihedral angles. Several clusters, derived from 3,0-helices and multipleturn conformations, had strong amino acid sequence patterns not evident among a-helices. Aspartate occurred over twice as frequently in the N-cap position of 310-helices as in the N-cap position of a-helices. Unlike a-helices, 310-helices had few C-termini ending in a left-handed a conformation; most 310 C-caps adopted an extended conformation. Differences in the distribution of hydrophobic residues among 3,0-and a-helices were also apparent, producing amphipathic 310-helices. Local interactions that stabilize 3,0-helices can be inferred both from the strong amino acid preferences found for these short helices, as well as from the existence of substructures in which tertiary interactions replace consensus local interactions. Because the folding and unfolding of a-helices have been postulated to proceed through reverse-turn and 310-helix intermediates, sequence differences between 310-and ahelices can also lend insight into factors influencing a-helix initiation and propagation. Keywords: a-helix; amino acid determinants; 310-helicesThe conformation of a segment of protein backbone is determined both by local interactions, i.e., interactions within the segment and with residues neighboring in sequence, and by interactions with the segment's tertiary environment. We have undertaken a series of studies of three-dimensional protein crystal structures, involving the systematic classification of local protein conformations and the analysis of correlations between amino acid sequence and local conformation classes (Karpen, 1991). Strong correlations between amino acid sequence and local conformation can point to side-chain interactions that stabilize or destabilize a particular conformation relative to other conformations. In this paper, we examine 310-helices and how their amino acid sequences differ from a-helices. The 310-helical structures are relatively common in proteins; 4% of all residues in our select data base of 51 high-resolution proteins are involved in 310-helices. These short helices usually occur on the surface of a pro-

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Section: Cluster 131mentioning

confidence: 99%

Differences in the amino acid distributions of 3₁₀‐helices and α‐helices

1992

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“…4 The enzyme active site is a heme group with a metal (iron) embedded in its center and that is linked to the protein via a sulfur bridge of a cysteinate residue (Cys 357 in P450 cam ). 5 The active center binds molecular oxygen, which is converted into an oxoiron(IV) active species, Compound I (Cpd I), via a series of reduction and protonation steps. Cpd I, however, is elusive, but kinetic isotope effects and product distributions implicate it to be involved in substrate monoxygenation reactions.…”

Section: Introductionmentioning

confidence: 99%

Effect of Porphyrin Ligands on the Regioselective Dehydrogenation versus Epoxidation of Olefins by Oxoiron(IV) Mimics of Cytochrome P450

et al. 2009

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The cytochromes P450 are versatile enzymes involved in various catalytic oxidation reactions, such as hydroxylation, epoxidation and dehydrogenation. In this work, we present combined experimental and theoretical studies on the change of regioselectivity in cyclohexadiene oxidation (i.e., epoxidation vs dehydrogenation) by oxoiron(IV) porphyrin complexes bearing different porphyrin ligands. Our experimental results show that meso-substitution of the porphyrin ring with electron-withdrawing substituents leads to a regioselectivity switch from dehydrogenation to epoxidation, affording the formation of epoxide as a major product. In contrast, electron-rich iron porphyrins are shown to produce benzene resulting from the dehydrogenation of cyclohexadiene. Density functional theory (DFT) calculations on the regioselectivity switch of epoxidation vs dehydrogenation have been performed using three oxoiron(IV) porphyrin oxidants with hydrogen atoms, phenyl groups, and pentachlorophenyl (ArCl 5 ) groups on the meso-position. The DFT studies show that the epoxidation reaction by the latter catalyst is stabilized because of favorable interactions of the substrate with halogen atoms of the meso-ligand as well as with pyrrole nitrogen atoms of the porphyrin macrocycle. Hydrogen abstraction transition states, in contrast, have a substrate-binding orientation further away from the porphyrin pyrrole nitrogens, and they are much less stabilized. Finally, the regioselectivity of dehydrogenation versus hydroxylation is rationalized using thermodynamic cycles.

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“…The decay of this intermediate involved a diffusion controlled protonation from its pH dependence and a deuterium isotope effect of 2. Lys 14 has been identified as a participating 2 nd sphere residue (mutation of this residue decreases the turnover rate by a factor of [20][21][22][23][24][25][26][27][28][29][30] and has been proposed to assist superoxide binding to the active site. 19,20 Though no intermediate has been trapped with the physiological substrate superoxide, Olivier et.…”

Section: Introductionmentioning

confidence: 99%

“…23 Alternatively, there is some similarity between the SOR active site and the active site of cytochrome P450 where the four porphyrin Nitrogens are in the equatorial plane and there is an axial cysteine ligand. 28 of an open co-ordination site has been suggested to be a possible reason for this 23 although the origin of the affinity for this 6 th ligand binding or the feasibility of a SOD reaction in a SOR active site has not been evaluated. Thus it is important to understand the role of the axial thiolate ligand and the lack of heme in determining the reactivity of SOR.…”

Section: Introductionmentioning

confidence: 99%

Sulfur K-Edge X-ray Absorption Spectroscopy and Density Functional Theory Calculations on Superoxide Reductase: Role of the Axial Thiolate in Reactivity

et al. 2007

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Superoxide reductase (SOR) is a non-heme iron enzyme which reduces superoxide to peroxide at a diffusion-controlled rate. Sulfur K-edge x-ray absorption spectroscopy (XAS) is used to investigate the ground state electronic structure of the resting high-spin and CN − bound low-spin Fe III forms of the 1Fe SOR from Pyrococcus furiosus. A computational model with constrained Imidazole rings (necessary for reproducing spin states), H-bonding interaction to the thiolate (necessary for reproducing Fe-S bond covalency of the high-spin and low-spin forms) and H-bonding to axial ligand (necessary to reproduce the ground state of the low-spin form) was developed and then used to investigate the enzymatic reaction mechanism. Reaction of the resting ferrous site with superoxide and protonation leading to a high-spin Fe III -OOH species and its subsequent protonation resulting in H 2 O 2 release is calculated to be the most energetically favorable reaction pathway. Our results suggest that the thiolate acts as a covalent anionic ligand. Replacing the thiolate with a neutral noncovalent ligand makes protonation very endothermic and greatly raises the reduction potential. The covalent nature of the thiolate weakens the Fe III bond to the proximal Oxygen of this hydroperoxo species, which raises its pK a by an additional 5 log units relative to a primarily anionic ligand facilitating its protonation. A comparison with Cytochrome P450 indicates that the stronger equatorial ligand field from the porphyrin results in a low-spin Fe III -OOH species which would not be capable of efficient H 2 O 2 release due to a spin-crossing barrier associated with formation of a high spin 5C Fe III product. Additionally, the presence of the di-anionic porphyrin π ring in Cytochrome P450 allows O-O heterolysis forming a Fe IV -oxo porphyrin radical species, which is calculated to be extremely unfavorable for the non-heme SOR ligand environment. Finally the 5C Fe III site which results from the product release at the end of the O 2 − reduction cycle is calculated to be capable of reacting with a second O 2 − resulting in superoxide dismutase (SOD) activity. However in contrast to FeSOD the 5C Fe III site of SOR which is more positive is calculated to have a high affinity for the binding a 6 th anionic ligand which would inhibit its SOD activity.

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High-resolution crystal structure of cytochrome P450cam

Cited by 1,363 publications

References 36 publications

Differences in the amino acid distributions of 3₁₀‐helices and α‐helices

Differences in the amino acid distributions of 3₁₀‐helices and α‐helices

Effect of Porphyrin Ligands on the Regioselective Dehydrogenation versus Epoxidation of Olefins by Oxoiron(IV) Mimics of Cytochrome P450

Sulfur K-Edge X-ray Absorption Spectroscopy and Density Functional Theory Calculations on Superoxide Reductase: Role of the Axial Thiolate in Reactivity

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High-resolution crystal structure of cytochrome P450cam

Cited by 1,363 publications

References 36 publications

Differences in the amino acid distributions of 310‐helices and α‐helices

Differences in the amino acid distributions of 310‐helices and α‐helices

Effect of Porphyrin Ligands on the Regioselective Dehydrogenation versus Epoxidation of Olefins by Oxoiron(IV) Mimics of Cytochrome P450

Sulfur K-Edge X-ray Absorption Spectroscopy and Density Functional Theory Calculations on Superoxide Reductase: Role of the Axial Thiolate in Reactivity

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Differences in the amino acid distributions of 3₁₀‐helices and α‐helices

Differences in the amino acid distributions of 3₁₀‐helices and α‐helices