• Home
  • Search Results
  • Iron(IV)hydroxide pK(a) and the role of thiolate ligation in C-H bond activation by cytochrome P450.

Iron(IV)hydroxide pK(a) and the role of thiolate ligation in C-H bond activation by cytochrome P450.

Science (New York, N.Y.) (2013-11-16)
Timothy H Yosca, Jonathan Rittle, Courtney M Krest, Elizabeth L Onderko, Alexey Silakov, Julio C Calixto, Rachel K Behan, Michael T Green
ABSTRACT

Cytochrome P450 enzymes activate oxygen at heme iron centers to oxidize relatively inert substrate carbon-hydrogen bonds. Cysteine thiolate coordination to iron is posited to increase the pK(a) (where K(a) is the acid dissociation constant) of compound II, an iron(IV)hydroxide complex, correspondingly lowering the one-electron reduction potential of compound I, the active catalytic intermediate, and decreasing the driving force for deleterious auto-oxidation of tyrosine and tryptophan residues in the enzyme's framework. Here, we report on the preparation of an iron(IV)hydroxide complex in a P450 enzyme (CYP158) in ≥90% yield. Using rapid mixing technologies in conjunction with Mössbauer, ultraviolet/visible, and x-ray absorption spectroscopies, we determine a pK(a) value for this compound of 11.9. Marcus theory analysis indicates that this elevated pK(a) results in a >10,000-fold reduction in the rate constant for oxidations of the protein framework, making these processes noncompetitive with substrate oxidation.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
L-Cysteine, 97%
Sigma-Aldrich
L-Tryptophan, reagent grade, ≥98% (HPLC)
Sigma-Aldrich
L-Cysteine, from non-animal source, BioReagent, suitable for cell culture, ≥98%
Sigma-Aldrich
L-Tyrosine, reagent grade, ≥98% (HPLC)
SAFC
L-Cysteine
Sigma-Aldrich
L-Tryptophan, from non-animal source, meets EP, JP, USP testing specifications, suitable for cell culture, 99.0-101.0%
Sigma-Aldrich
L-Cysteine, BioUltra, ≥98.5% (RT)
Sigma-Aldrich
L-Tryptophan, BioUltra, ≥99.5% (NT)
Sigma-Aldrich
L-Tyrosine, from non-animal source, meets EP, USP testing specifications, suitable for cell culture, 99.0-101.0%
Supelco
Tryptophan, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
L-Cysteine, ≥97%, FG
Sigma-Aldrich
L-Tyrosine, BioUltra, ≥99.0% (NT)
Supelco
L-Tryptophan, certified reference material, TraceCERT®
Supelco
L-Tyrosine, Pharmaceutical Secondary Standard; Certified Reference Material
SAFC
L-Tyrosine
Supelco
L-Cysteine, certified reference material, TraceCERT®
Sigma-Aldrich
L-Cysteine, produced by Wacker Chemie AG, Burghausen, Germany, ≥98.0%
Sigma-Aldrich
L-Tyrosine, FG
Supelco
L-Tyrosine, certified reference material, TraceCERT®