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In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant.
Cohen, Michal; Pignatti, Emanuele; Dines, Monica; Mory, Adi; Ekhilevitch, Nina; Kolodny, Rachel; Flück, Christa E; Tiosano, Dov.
Afiliación
  • Cohen M; Pediatric Endocrinology Unit, Ruth Rappaport Children's Hospital, Rambam Healthcare Campus, Haifa 352540, Israel.
  • Pignatti E; The Ruth and Bruce Rappaport Faculty of Medicine, Technion, Israel Institute of Technology, Haifa 352540, Israel.
  • Dines M; Pediatric Endocrinology, Diabetology and Metabolism, Department of Pediatrics, University Hospital Inselspital, University of Bern, 3010 Bern, Switzerland.
  • Mory A; Department of BioMedical Research, University Hospital Inselspital, University of Bern, 3010 Bern, Switzerland.
  • Ekhilevitch N; Sagol Department of Neurobiology, University of Haifa, Mount Carmel, Haifa 31905, Israel.
  • Kolodny R; Genetics Institute, Rambam Health Care Campus, Haifa 3525408, Israel.
  • Flück CE; Genetics Institute, Rambam Health Care Campus, Haifa 3525408, Israel.
  • Tiosano D; Department of Computer Science, University of Haifa, Mount Carmel, Haifa 3498838, Israel.
Int J Mol Sci ; 21(16)2020 Aug 14.
Article en En | MEDLINE | ID: mdl-32824094
Classical congenital adrenal hyperplasia (CAH) caused by pathogenic variants in the steroid 21-hydroxylase gene (CYP21A2) is a severe life-threatening condition. We present a detailed investigation of the molecular and functional characteristics of a novel pathogenic variant in this gene. The patient, 46 XX newborn, was diagnosed with classical salt wasting CAH in the neonatal period after initially presenting with ambiguous genitalia. Multiplex ligation-dependent probe analysis demonstrated a full deletion of the paternal CYP21A2 gene, and Sanger sequencing revealed a novel de novo CYP21A2 variant c.694-696del (E232del) in the other allele. This variant resulted in the deletion of a non-conserved single amino acid, and its functional relevance was initially undetermined. We used both in silico and in vitro methods to determine the mechanistic significance of this mutation. Computational analysis relied on the solved structure of the protein (Protein-data-bank ID 4Y8W), structure prediction of the mutated protein, evolutionary analysis, and manual inspection. We predicted impaired stability and functionality of the protein due to a rotatory disposition of amino acids in positions downstream of the deletion. In vitro biochemical evaluation of enzymatic activity supported these predictions, demonstrating reduced protein levels to 22% compared to the wild-type form and decreased hydroxylase activity to 1-4%. This case demonstrates the potential of combining in-silico analysis based on evolutionary information and structure prediction with biochemical studies. This approach can be used to investigate other genetic variants to understand their potential effects.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Simulación por Computador / Esteroide 21-Hidroxilasa / Mutación Tipo de estudio: Guideline / Prognostic_studies Límite: Child, preschool / Female / Humans / Infant / Newborn Idioma: En Revista: Int J Mol Sci Año: 2020 Tipo del documento: Article País de afiliación: Israel Pais de publicación: Suiza
Texto completo
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Simulación por Computador / Esteroide 21-Hidroxilasa / Mutación Tipo de estudio: Guideline / Prognostic_studies Límite: Child, preschool / Female / Humans / Infant / Newborn Idioma: En Revista: Int J Mol Sci Año: 2020 Tipo del documento: Article País de afiliación: Israel Pais de publicación: Suiza