D-2-hydroxyglutaric aciduria Type I: Functional analysis of D2HGDH missense variants

Ana Pop, Eduard A. Struys, Erwin E. W. Jansen, Matilde R. Fernandez, Warsha A. Kanhai, Silvy J. M. van Dooren, Senay Ozturk, Justin van Oostendorp, Pascal Lennertz, Martijn Kranendijk, Marjo S. van der Knaap, K. Michael Gibson, Emile van Schaftingen, Gajja S. Salomons

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

D-2-hydroxyglutaric aciduria Type I (D-2-HGA Type I), a neurometabolic disorder with a broad clinical spectrum, is caused by recessive variants in the D2HGDH gene encoding D-2-hydroxyglutarate dehydrogenase (D-2-HGDH). We and others detected 42 potentially pathogenic variants in D2HGDH of which 31 were missense. We developed functional studies to investigate the effect of missense variants on D-2-HGDH catalytic activity. Site-directed mutagenesis was used to introduce 31 missense variants in the pCMV5-D2HGDH expression vector. The wild type and missense variants were overexpressed in HEK293 cells. D-2-HGDH enzyme activity was evaluated based on the conversion of [ 2 H 4 ]D-2-HG to [ 2 H 4 ]2-ketoglutarate, which was subsequently converted into [ 2 H 4 ]L-glutamate and the latter quantified by LC-MS/MS. Eighteen variants resulted in almost complete ablation of D-2-HGDH activity and thus, should be considered pathogenic. The remaining 13 variants manifested residual activities ranging between 17% and 94% of control enzymatic activity. Our functional assay evaluating the effect of novel D2HGDH variants will be beneficial for the classification of missense variants and determination of pathogenicity.
Original languageEnglish
JournalHuman Mutation
DOIs
Publication statusPublished - 2019

Cite this

Pop, Ana ; Struys, Eduard A. ; Jansen, Erwin E. W. ; Fernandez, Matilde R. ; Kanhai, Warsha A. ; van Dooren, Silvy J. M. ; Ozturk, Senay ; van Oostendorp, Justin ; Lennertz, Pascal ; Kranendijk, Martijn ; van der Knaap, Marjo S. ; Gibson, K. Michael ; van Schaftingen, Emile ; Salomons, Gajja S. / D-2-hydroxyglutaric aciduria Type I: Functional analysis of D2HGDH missense variants. In: Human Mutation. 2019.
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title = "D-2-hydroxyglutaric aciduria Type I: Functional analysis of D2HGDH missense variants",
abstract = "D-2-hydroxyglutaric aciduria Type I (D-2-HGA Type I), a neurometabolic disorder with a broad clinical spectrum, is caused by recessive variants in the D2HGDH gene encoding D-2-hydroxyglutarate dehydrogenase (D-2-HGDH). We and others detected 42 potentially pathogenic variants in D2HGDH of which 31 were missense. We developed functional studies to investigate the effect of missense variants on D-2-HGDH catalytic activity. Site-directed mutagenesis was used to introduce 31 missense variants in the pCMV5-D2HGDH expression vector. The wild type and missense variants were overexpressed in HEK293 cells. D-2-HGDH enzyme activity was evaluated based on the conversion of [ 2 H 4 ]D-2-HG to [ 2 H 4 ]2-ketoglutarate, which was subsequently converted into [ 2 H 4 ]L-glutamate and the latter quantified by LC-MS/MS. Eighteen variants resulted in almost complete ablation of D-2-HGDH activity and thus, should be considered pathogenic. The remaining 13 variants manifested residual activities ranging between 17{\%} and 94{\%} of control enzymatic activity. Our functional assay evaluating the effect of novel D2HGDH variants will be beneficial for the classification of missense variants and determination of pathogenicity.",
author = "Ana Pop and Struys, {Eduard A.} and Jansen, {Erwin E. W.} and Fernandez, {Matilde R.} and Kanhai, {Warsha A.} and {van Dooren}, {Silvy J. M.} and Senay Ozturk and {van Oostendorp}, Justin and Pascal Lennertz and Martijn Kranendijk and {van der Knaap}, {Marjo S.} and Gibson, {K. Michael} and {van Schaftingen}, Emile and Salomons, {Gajja S.}",
year = "2019",
doi = "10.1002/humu.23751",
language = "English",
journal = "Human Mutation",
issn = "1059-7794",
publisher = "Wiley-Liss Inc.",

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D-2-hydroxyglutaric aciduria Type I: Functional analysis of D2HGDH missense variants. / Pop, Ana; Struys, Eduard A.; Jansen, Erwin E. W.; Fernandez, Matilde R.; Kanhai, Warsha A.; van Dooren, Silvy J. M.; Ozturk, Senay; van Oostendorp, Justin; Lennertz, Pascal; Kranendijk, Martijn; van der Knaap, Marjo S.; Gibson, K. Michael; van Schaftingen, Emile; Salomons, Gajja S.

In: Human Mutation, 2019.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - D-2-hydroxyglutaric aciduria Type I: Functional analysis of D2HGDH missense variants

AU - Pop, Ana

AU - Struys, Eduard A.

AU - Jansen, Erwin E. W.

AU - Fernandez, Matilde R.

AU - Kanhai, Warsha A.

AU - van Dooren, Silvy J. M.

AU - Ozturk, Senay

AU - van Oostendorp, Justin

AU - Lennertz, Pascal

AU - Kranendijk, Martijn

AU - van der Knaap, Marjo S.

AU - Gibson, K. Michael

AU - van Schaftingen, Emile

AU - Salomons, Gajja S.

PY - 2019

Y1 - 2019

N2 - D-2-hydroxyglutaric aciduria Type I (D-2-HGA Type I), a neurometabolic disorder with a broad clinical spectrum, is caused by recessive variants in the D2HGDH gene encoding D-2-hydroxyglutarate dehydrogenase (D-2-HGDH). We and others detected 42 potentially pathogenic variants in D2HGDH of which 31 were missense. We developed functional studies to investigate the effect of missense variants on D-2-HGDH catalytic activity. Site-directed mutagenesis was used to introduce 31 missense variants in the pCMV5-D2HGDH expression vector. The wild type and missense variants were overexpressed in HEK293 cells. D-2-HGDH enzyme activity was evaluated based on the conversion of [ 2 H 4 ]D-2-HG to [ 2 H 4 ]2-ketoglutarate, which was subsequently converted into [ 2 H 4 ]L-glutamate and the latter quantified by LC-MS/MS. Eighteen variants resulted in almost complete ablation of D-2-HGDH activity and thus, should be considered pathogenic. The remaining 13 variants manifested residual activities ranging between 17% and 94% of control enzymatic activity. Our functional assay evaluating the effect of novel D2HGDH variants will be beneficial for the classification of missense variants and determination of pathogenicity.

AB - D-2-hydroxyglutaric aciduria Type I (D-2-HGA Type I), a neurometabolic disorder with a broad clinical spectrum, is caused by recessive variants in the D2HGDH gene encoding D-2-hydroxyglutarate dehydrogenase (D-2-HGDH). We and others detected 42 potentially pathogenic variants in D2HGDH of which 31 were missense. We developed functional studies to investigate the effect of missense variants on D-2-HGDH catalytic activity. Site-directed mutagenesis was used to introduce 31 missense variants in the pCMV5-D2HGDH expression vector. The wild type and missense variants were overexpressed in HEK293 cells. D-2-HGDH enzyme activity was evaluated based on the conversion of [ 2 H 4 ]D-2-HG to [ 2 H 4 ]2-ketoglutarate, which was subsequently converted into [ 2 H 4 ]L-glutamate and the latter quantified by LC-MS/MS. Eighteen variants resulted in almost complete ablation of D-2-HGDH activity and thus, should be considered pathogenic. The remaining 13 variants manifested residual activities ranging between 17% and 94% of control enzymatic activity. Our functional assay evaluating the effect of novel D2HGDH variants will be beneficial for the classification of missense variants and determination of pathogenicity.

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UR - https://www.ncbi.nlm.nih.gov/pubmed/30908763

U2 - 10.1002/humu.23751

DO - 10.1002/humu.23751

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JO - Human Mutation

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SN - 1059-7794

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