Heterozygosity for an in-frame deletion causes glutaryl-CoA dehydrogenase deficiency in a patient detected by newborn screening: investigation of the effect of the mutant allele
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Heterozygosity for an in-frame deletion causes glutaryl-CoA dehydrogenase deficiency in a patient detected by newborn screening : investigation of the effect of the mutant allele. / Bross, Peter Gerd; Frederiksen, Jane B; Bie, Anne Sigaard; Hansen, Jakob; Palmfeldt, Johan; Nielsen, Marit N; Duno, Morten; Lund, Allan M; Christensen, Ernst.
I: Journal of Inherited Metabolic Disease, Bind 35, Nr. 5, 2012, s. 787-96.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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TY - JOUR
T1 - Heterozygosity for an in-frame deletion causes glutaryl-CoA dehydrogenase deficiency in a patient detected by newborn screening
T2 - investigation of the effect of the mutant allele
AU - Bross, Peter Gerd
AU - Frederiksen, Jane B
AU - Bie, Anne Sigaard
AU - Hansen, Jakob
AU - Palmfeldt, Johan
AU - Nielsen, Marit N
AU - Duno, Morten
AU - Lund, Allan M
AU - Christensen, Ernst
PY - 2012
Y1 - 2012
N2 - A patient with suspected glutaric aciduria type 1 (GA-1) was detected by newborn screening. GA-1 is known as an autosomal recessively inherited disease due to defects in the gene coding for glutaryl-CoA dehydrogenase (GCDH), a mitochondrial enzyme involved in the catabolism of the amino acids hydroxylysine, lysine and tryptophan. DNA and cDNA sequencing revealed a 18 bp deletion (c.553_570del18) resulting in deletion of six amino acids (p.Gly185_Ser190del) in one allele and no sequence changes in the other allele. Confirmatory biochemical analysis of blood, urine and cultured fibroblasts from the proband were consistent with a mild biochemical GA-1 phenotype. Recombinant expression of the mutant variant in E. coli showed that the GCDH-(p.Gly185_Ser190del) protein displayed severely decreased assembly into tetramers and enzyme activity. To discover a potential dominant negative effect of the mutant protein, we engineered a prokaryotic expression system in which expression of a wild type and a mutant GCDH allele is controlled by separately inducible promoters. These cells displayed decreased levels of GCDH tetramer and enzyme activity when expressing both the wild type and the mutant GCDH variant protein compared to the situation when only the wild type allele was expressed. Further experiments suggest that the major impact of the GCDH-(p.Gly185_Ser190del) protein in heterozygous cells consists of hampering the assembly of wild type GCDH into tetramers. Our experimental data are consistent with the hypothesis that heterozygosity for this mutation confers a dominant negative effect resulting in a GCDH enzyme activity that is significantly lower than the expected 50%.
AB - A patient with suspected glutaric aciduria type 1 (GA-1) was detected by newborn screening. GA-1 is known as an autosomal recessively inherited disease due to defects in the gene coding for glutaryl-CoA dehydrogenase (GCDH), a mitochondrial enzyme involved in the catabolism of the amino acids hydroxylysine, lysine and tryptophan. DNA and cDNA sequencing revealed a 18 bp deletion (c.553_570del18) resulting in deletion of six amino acids (p.Gly185_Ser190del) in one allele and no sequence changes in the other allele. Confirmatory biochemical analysis of blood, urine and cultured fibroblasts from the proband were consistent with a mild biochemical GA-1 phenotype. Recombinant expression of the mutant variant in E. coli showed that the GCDH-(p.Gly185_Ser190del) protein displayed severely decreased assembly into tetramers and enzyme activity. To discover a potential dominant negative effect of the mutant protein, we engineered a prokaryotic expression system in which expression of a wild type and a mutant GCDH allele is controlled by separately inducible promoters. These cells displayed decreased levels of GCDH tetramer and enzyme activity when expressing both the wild type and the mutant GCDH variant protein compared to the situation when only the wild type allele was expressed. Further experiments suggest that the major impact of the GCDH-(p.Gly185_Ser190del) protein in heterozygous cells consists of hampering the assembly of wild type GCDH into tetramers. Our experimental data are consistent with the hypothesis that heterozygosity for this mutation confers a dominant negative effect resulting in a GCDH enzyme activity that is significantly lower than the expected 50%.
U2 - 10.1007/s10545-011-9437-y
DO - 10.1007/s10545-011-9437-y
M3 - Journal article
C2 - 22231382
VL - 35
SP - 787
EP - 796
JO - Journal of Inherited Metabolic Disease
JF - Journal of Inherited Metabolic Disease
SN - 0141-8955
IS - 5
ER -
ID: 48516675