Functional characterization of three CYP21A2 sequence variants (p.A265V, p.W302S, p.D322G) employing a yeast co-expression system.
Congenital adrenal hyperplasia (CAH) due to steroid 21-hydroxylase (CYP21A2) deficiency is the commonest inborn error in steroid hormone biosynthesis. Functional in vitro assessment of mutant activity generally correlates well with clinical phenotype and therefore has contributed greatly to phenotype prediction in this CAH variant. Three CYP21A2 sequence variants (g.1641C>T, p.A265V; g.1752G>C, p.W302S; and g.2012A>G, p.D322G) identified in patients with non-classic and simple virilizing CAH were characterized using a yeast co-expression system and a computational three-dimensional CYP21A2 model. Computational analysis of the mutants in the three-dimensional structural model predicted no relevant effect of p.A265V, while p.W302S and p.D322G were predicted to impact significantly on enzyme function. Consistent with these findings, in vitro mutant analysis revealed enzyme activity similar to wild-type for p.A265V, whereas p.W302S and p.D322G exerted activities compatible with simple virilizing and non-classical CAH, respectively. The results indicate that p.A265V is an allelic variant rather than a disease-causing amino acid change, whilst p.W302S and p.D322G could be confirmed as functionally relevant mutations. These findings emphasize the value of in vitro functional analysis of sequence variations in predicting genotype-phenotype correlations and disease severity.
Bleicken
Caroline
C
Institute of Biomedical Research, Division of Medical Sciences, University of Birmingham, Birmingham, United Kingdom.
Loidi
Lourdes
L
Dhir
Vivek
V
Parajes
Silvia
S
Quinteiro
Celsa
C
Dominguez
Fernando
F
Grötzinger
Joachim
J
Sippell
Wolfgang G
WG
Riepe
Felix G
FG
Arlt
Wiebke
W
Krone
Nils
N
D000293:Adolescent; D002648:Child; D005260:Female; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D050505:Mutant Proteins / Q000378:metabolism*; D009154:Mutation / Q000235:genetics*; D014176:Protein Biosynthesis; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D013255:Steroid 21-Hydroxylase / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic
D050505:Mutant Proteins; C531510:CYP21A2 protein, human; D013255:Steroid 21-Hydroxylase