© 1994 Oxford University Press
OTHER |
Functional characterization of the novel L108W and P186L mutations detected in the type II 3ß-hydroxysteroid dehydrogenase gene of a male pseudohermaphrodite with congenital adrenal hyperplasia
Medical Research Council Group in Molecular Endocrinology, CHUL Research Center and Laval University 2705 Laurier Boulevard, Québec G1V 4G2, Canada
1INSERM U329 and Department of Pediatrics, Université de Lyon and H
pital Debrousse, Lyon Cedex 05, France
*To whom correspondence should be addressed
Received May 25, 1994; Revised July 14, 1994; Accepted July 14, 1994
Two Isoenzymes are responsible for 3ß-hydroxysterold dehydrogenase/
5-4-isomerase (3ß-HSD) activity in humans. We analyzed the structure of types I and II 3ß-HSD genes in a male pseudohermaphrodite suffering from a severe salt-losing form of congenital adrenal hyperplasia. We did not detect any mutation in the type I 3ß-HSD gene, but we found two different missense mutations in exon IV of the type II 3ß-HSD gene of the patient; a conversion of codon Leu108 into a Trp (L108W) inherited from his mother and a conversion of codon Pro186 into a Leu (P186L) inherited from his father. We assessed the effect of the L108W and P186L mutations on 3ß-HSD activity by in vitro analysis of mutant enzymes expressed in heterologous COS-1 cells. Using homogenates from transfected cells, the Km values for PREG were 7 ± 2 and 8 ± 2 µM for the recombinant L108W and P186L enzymes, respectively, compared with 2.2 ± 0.2 µM for the normal type II 3ß-HSD enzyme. Moreover, Km values for NAD+ were much higher for the L108W and P186L proteins, being 678 ± 166 and 920 ± 351 µM, respectively, compared with 24 ± 3 µM for the normal type II 3ß-HSD enzyme. Vmax values for PREG and NAD+ were lower for both mutant enzymes; thus, the in vitro overall efficiency, relative to the normal enzyme, is approximate as 0.3% and 0.2% for the L108W and P186L enzymes, respectively. The present study is the first description of mutations which significantly affect the affinity for NAD+ in addition to reducing the affinity for PREG, thus providing useful information on the structure-activity relationships of the 3ß-HSD enzyme superfamily. Moreover, this patient is the first case presenting the salt-wasting form of classical 3ß-HSD deficiency caused by mutated alleles possessing residual enzymatic activity. The combination of decreased Km values for the sterold substrate and cofactor thus explains the severe form of this enzymatic defect responsible for congenital adrenal hyperplasia and male pseudohermaphroditlsm.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  J. Simard, M.-L. Ricketts, S. Gingras, P. Soucy, F. A. Feltus, and M. H. Melner Molecular Biology of the 3{beta}-Hydroxysteroid Dehydrogenase/{Delta}5-{Delta}4 Isomerase Gene Family Endocr. Rev., June 1, 2005; 26(4): 525 - 582. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. H. Johannsen, D. Mallet, H. Dige-Petersen, J. Muller, K. M. Main, Y. Morel, and M. G. Forest Delayed Diagnosis of Congenital Adrenal Hyperplasia with Salt Wasting Due to Type II 3{beta}-Hydroxysteroid Dehydrogenase Deficiency J. Clin. Endocrinol. Metab., April 1, 2005; 90(4): 2076 - 2080. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Zhang, J. I. Mason, Y. Naiki, K. C. Copeland, M. Castro-Magana, T. T. Gordon-Walker, Y. T. Chang, and S. Pang Characterization of Two Novel Homozygous Missense Mutations Involving Codon 6 and 259 of Type II 3{beta}-Hydroxysteroid Dehydrogenase (3{beta}HSD) Gene Causing, Respectively, Nonsalt-Wasting and Salt-Wasting 3{beta}HSD Deficiency Disorder J. Clin. Endocrinol. Metab., April 1, 2000; 85(4): 1678 - 1685. [Abstract] [Full Text]
|
|