5VBU

Crystal Structure of Human Cytochrome P450 21A2 Hydroxyprogesterone Complex

5VBU の概要

• エントリーDOI: 10.2210/pdb5vbu/pdb
• 分子名称: Cytochrome P450 21-hydroxylase, PROTOPORPHYRIN IX CONTAINING FE, (9beta)-17-hydroxypregn-4-ene-3,20-dione, ... (4 entities in total)
• 機能のキーワード: steroid hydroxylation, monooxygenases, adrenal steroidogenesis, congenital adrenal hyperplasia, addison's disease, kinetic isotope effects, oxidoreductase, hydroxyprogesterone
• 由来する生物種: Homo sapiens (Human)
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 163971.72

構造登録者

Pallan, P.S.,Egli, M. (登録日: 2017-03-30, 公開日: 2017-05-31, 最終更新日: 2024-05-22)

主引用文献

Wang, C.,Pallan, P.S.,Zhang, W.,Lei, L.,Yoshimoto, F.K.,Waterman, M.R.,Egli, M.,Guengerich, F.P.
Functional analysis of human cytochrome P450 21A2 variants involved in congenital adrenal hyperplasia.
J. Biol. Chem., 292:10767-10778, 2017

PubMed Abstract: Cytochrome P450 (P450, CYP) 21A2 is the major steroid 21-hydroxylase, converting progesterone to 11-deoxycorticosterone and 17α-hydroxyprogesterone (17α-OH-progesterone) to 11-deoxycortisol. More than 100 variants give rise to congenital adrenal hyperplasia (CAH). We previously reported a structure of WT human P450 21A2 with bound progesterone and now present a structure bound to the other substrate (17α-OH-progesterone). We found that the 17α-OH-progesterone- and progesterone-bound complex structures are highly similar, with only some minor differences in surface loop regions. Twelve P450 21A2 variants associated with either salt-wasting or nonclassical forms of CAH were expressed, purified, and analyzed. The catalytic activities of these 12 variants ranged from 0.00009% to 30% of WT P450 21A2 and the extent of heme incorporation from 10% to 95% of the WT. Substrate dissociation constants () for four variants were 37-13,000-fold higher than for WT P450 21A2. Cytochrome , which augments several P450 activities, inhibited P450 21A2 activity. Similar to the WT enzyme, high noncompetitive intermolecular kinetic deuterium isotope effects (≥ 5.5) were observed for all six P450 21A2 variants examined for 21-hydroxylation of 21--progesterone, indicating that C-H bond breaking is a rate-limiting step over a 10-fold range of catalytic efficiency. Using UV-visible and CD spectroscopy, we found that P450 21A2 thermal stability assessed in bacterial cells and with purified enzymes differed among salt-wasting- and nonclassical-associated variants, but these differences did not correlate with catalytic activity. Our in-depth investigation of CAH-associated P450 21A2 variants reveals critical insight into the effects of disease-causing mutations on this important enzyme.

PubMed: 28539365
DOI: 10.1074/jbc.M117.792465

実験手法

X-RAY DIFFRACTION (3.31 Å)