Functional characterization of eight rare missense CYP1B1 variants involved in congenital glaucoma and their association with null genotypes
Purpose: To evaluate the function of eight missense CYP1B1 single nucleotide variants(SNVs) previously identified in patients with primary congenital glaucoma (PCG).Methods: The eight variants were obtained by site-directed mutagenesis andtransiently expressed in human embryonic kidney 293-T (HEK-293...
| Autores: | , , , , |
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| Tipo de recurso: | artículo |
| Fecha de publicación: | 2016 |
| País: | España |
| Institución: | Universidad Católica de Valencia San Vicente Mártir |
| Repositorio: | RIUCV. Repositorio de la Universidad Católica de Valencia San Vicente Mártir |
| Idioma: | inglés |
| OAI Identifier: | oai:riucv.ucv.es:20.500.12466/5047 |
| Acceso en línea: | http://hdl.handle.net/20.500.12466/5047 |
| Access Level: | acceso abierto |
| Palabra clave: | CYP1B1 Primary congenital glaucoma 2302 Bioquímica |
| Sumario: | Purpose: To evaluate the function of eight missense CYP1B1 single nucleotide variants(SNVs) previously identified in patients with primary congenital glaucoma (PCG).Methods: The eight variants were obtained by site-directed mutagenesis andtransiently expressed in human embryonic kidney 293-T (HEK-293T) cells. Thecatalytic activity, protein stability and subcellular localization of the differentrecombinant CYP1B1 variants were assessed in this cell line.Results: Six of the mutant CYP1B1 proteins (p.L89P, p.A106D, p.R390S,p.P437L, p.C470Y and S485F) showed catalytic activity values ranging from0% to 4% of those of the wild-type protein and were considered null variants.The activity values of the two remaining variants (p.F123L and p.A237E) wereclose to 20% of that of the wild-type enzyme and were classified as hypomorphicvariants. Reduced protein stability contributed partially to the decreasedcatalytic activity of two of the mutant enzymes (p.L89P and p.A106D). Noneof the CYP1B1 variants showed intracellular aggregation and they all displayeda normal subcellular localization in the endoplasmic reticulum, suggesting thatthey had folded into a wild-type-like structure. The enzymatic activity associatedwith the different genotypes in which these CYP1B1 variants were present wasestimated to range from 0% to 10% of that of the wild-type genotype.Conclusion: These results confirm the pathogenicity of the analysed missenseCYP1B1 variants and further support the concept that either absent or very lowCYP1B1 activity levels are the primary molecular defect involved in PCGpathogenesis. |
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