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In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant
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 patien...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7461554/ https://www.ncbi.nlm.nih.gov/pubmed/32824094 http://dx.doi.org/10.3390/ijms21165857 |
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author | Cohen, Michal Pignatti, Emanuele Dines, Monica Mory, Adi Ekhilevitch, Nina Kolodny, Rachel Flück, Christa E. Tiosano, Dov |
author_facet | Cohen, Michal Pignatti, Emanuele Dines, Monica Mory, Adi Ekhilevitch, Nina Kolodny, Rachel Flück, Christa E. Tiosano, Dov |
author_sort | Cohen, Michal |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-7461554 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-74615542020-09-04 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 Int J Mol Sci Communication 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. MDPI 2020-08-14 /pmc/articles/PMC7461554/ /pubmed/32824094 http://dx.doi.org/10.3390/ijms21165857 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Communication Cohen, Michal Pignatti, Emanuele Dines, Monica Mory, Adi Ekhilevitch, Nina Kolodny, Rachel Flück, Christa E. Tiosano, Dov In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant |
title | In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant |
title_full | In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant |
title_fullStr | In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant |
title_full_unstemmed | In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant |
title_short | In Silico Structural and Biochemical Functional Analysis of a Novel CYP21A2 Pathogenic Variant |
title_sort | in silico structural and biochemical functional analysis of a novel cyp21a2 pathogenic variant |
topic | Communication |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7461554/ https://www.ncbi.nlm.nih.gov/pubmed/32824094 http://dx.doi.org/10.3390/ijms21165857 |
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