3BE0

The Role of Asn 242 in P450cin

3BE0 の概要

• エントリーDOI: 10.2210/pdb3be0/pdb
• 関連するPDBエントリー: 1T2B 3BDZ
• 分子名称: P450cin, PROTOPORPHYRIN IX CONTAINING FE, 1,3,3-TRIMETHYL-2-OXABICYCLO[2.2.2]OCTANE, ... (4 entities in total)
• 機能のキーワード: protein substrate complex, unknown function
• 由来する生物種: Citrobacter braakii
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 90768.79

構造登録者

Meharenna, Y.T.,Poulos, T.L. (登録日: 2007-11-15, 公開日: 2008-02-12, 最終更新日: 2025-04-30)

主引用文献

Meharenna, Y.T.,Slessor, K.E.,Cavaignac, S.M.,Poulos, T.L.,De Voss, J.J.
The critical role of substrate-protein hydrogen bonding in the control of regioselective hydroxylation in p450cin
J.Biol.Chem., 283:10804-10812, 2008

PubMed Abstract: Cytochrome P450cin (CYP176A1) is a bacterial P450 isolated from Citrobacter braakii that catalyzes the hydroxylation of cineole to (S)-6beta-hydroxycineole. This initiates the biodegradation of cineole, enabling C. braakii to live on cineole as its sole source of carbon and energy. P450cin lacks the almost universally conserved threonine residue believed to be involved in dioxygen activation and instead contains an asparagine at this position (Asn-242). To investigate the role of Asn-242 in P450cin catalysis, it was converted to alanine, and the resultant mutant was characterized. The characteristic CO-bound spectrum and spectrally determined K(D) for substrate binding were unchanged in the mutant. The x-ray crystal structures of the substrate-free and -bound N242A mutant were determined and show that the only significant change is in a reorientation of the substrate such that (R)-6alpha-hydroxycineole should be a major product. Molecular dynamics simulations of both wild type and mutant are consistent with the change in regio- and stereoselectivity predicted from the crystal structure. The mutation has only a modest effect on enzyme activity and on the diversion of the NADPH-reducing equivalent toward unproductive peroxide formation. Product profile analysis shows that (R)-6alpha-hydroxycineole is the main product, which is consistent with the crystal structure. These results demonstrate that Asn-242 is not a functional replacement for the conserved threonine in other P450s but, rather, is critical in controlling regioselective substrate oxidation.

PubMed: 18270198
DOI: 10.1074/jbc.M709722200

実験手法

X-RAY DIFFRACTION (3.05 Å)