TY - JOUR
T1 - Mutations of P450c21 (steroid 21-hydroxylase) at Cys428, Val281, and Ser268 result in complete, partial, or no loss of enzymatic activity, respectively
AU - Wu, Du An
AU - Chung, Bon Chu
N1 - Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 1991/8
Y1 - 1991/8
N2 - Steroid 21-hydroxylase deficiency is the major cause of congenital adrenal hyperplasia (CAH), a common genetic disease. To define the relationship between gene mutations and enzyme deficiency, we generated missense mutations of the 21-hydroxylase cDNA at three different sites and characterized the mutant proteins after expressing them in cultured mammalian and yeast cells. Among them, Ser268 and Val281 have been found to be mutated in CAH patients, whereas Cys428 has been implicated as the heme ligand. Our results show mutations at these sites result in complete, partial, or no loss of the enzymatic activity. All the Cys428 mutants had neither enzymatic activity nor P450 absorption, thus supporting the notion that Cys428 is the heme ligand. All the 268-mutants exhibited the same activity as normal 21-hydroxylase, demonstrating that the clinically observed Ser268 → Thr change represents a polymorphism rather than the cause of the enzyme deficiency. The 281-mutants had normal Km but greatly reduced Vmax values that also paralleled the reduction in the heme content, in the order Val281 (normal, 100%) > Ile281 (50%) > Leu281 (20%) > Thr281 (10%). Our findings suggest that the methyl group at the β-carbon of Val281 is required for heme incorporation and consequently enzymatic activity.
AB - Steroid 21-hydroxylase deficiency is the major cause of congenital adrenal hyperplasia (CAH), a common genetic disease. To define the relationship between gene mutations and enzyme deficiency, we generated missense mutations of the 21-hydroxylase cDNA at three different sites and characterized the mutant proteins after expressing them in cultured mammalian and yeast cells. Among them, Ser268 and Val281 have been found to be mutated in CAH patients, whereas Cys428 has been implicated as the heme ligand. Our results show mutations at these sites result in complete, partial, or no loss of the enzymatic activity. All the Cys428 mutants had neither enzymatic activity nor P450 absorption, thus supporting the notion that Cys428 is the heme ligand. All the 268-mutants exhibited the same activity as normal 21-hydroxylase, demonstrating that the clinically observed Ser268 → Thr change represents a polymorphism rather than the cause of the enzyme deficiency. The 281-mutants had normal Km but greatly reduced Vmax values that also paralleled the reduction in the heme content, in the order Val281 (normal, 100%) > Ile281 (50%) > Leu281 (20%) > Thr281 (10%). Our findings suggest that the methyl group at the β-carbon of Val281 is required for heme incorporation and consequently enzymatic activity.
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U2 - 10.1172/JCI115334
DO - 10.1172/JCI115334
M3 - Article
C2 - 1864962
AN - SCOPUS:0025772912
VL - 88
SP - 519
EP - 523
JO - Journal of Clinical Investigation
JF - Journal of Clinical Investigation
SN - 0021-9738
IS - 2
ER -