Detection of heterozygotes in maple-syrup-urine disease: Measurements of branched-chain α-ketoacid dehydrogenase and its components in cell cultures

D. T. Chuang, L. S. Ku, D. S. Kerr, R. P. Cox

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

To detect heterozygotes for maple-syrup-urine disease (MSUD), activities of branched-chain α-ketoacid (BCKA) dehydrogenase and its components in skin fibroblasts of two obligatory heterozygotes and amnion cells of a fetus at risk were measured. Intact heterozygous cells were found to decarboxylate [1-14C]α-ketoisovalerate at rates equal to or only slightly lower than normal subjects. The inability to differentiate heterozygotes from normals with the intact cell assay confirms earlier studies with intact leukocytes using [1-14C]leucine as a substrate. By contrast, measurements of BCKA dehydrogenase activity with disrupted cell suspensions showed MSUD heterozygotes with 30%-60% of normal activity. Moreover, biphasic kinetics in heterozygous cells were observed with increasing substrate concentrations. The altered biphasic kinetics probably reflect expression of the normal allele in the early hyperbolic portion of the curve and of the mutant allele in the later secondary rise at high substrate concentrations. Assays of component activities showed concordant E1 decarboxylase deficiency in both heterozygous- and homozygous-affected cells, whereas the E3, dihydrolipoyl dehydrogenase-component, activity was normal. The above results taken together appear to provide an approach to detection of the heterozygote in MSUD.

Original languageEnglish (US)
Pages (from-to)416-424
Number of pages9
JournalAmerican Journal of Human Genetics
Volume34
Issue number3
StatePublished - 1982

Fingerprint

Heterozygote Detection
3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)
Maple Syrup Urine Disease
Cell Culture Techniques
Heterozygote
Alleles
Dihydrolipoamide Dehydrogenase
Amnion
Carboxy-Lyases
Leucine
Suspensions
Fetus
Leukocytes
Fibroblasts
Skin

ASJC Scopus subject areas

  • Genetics

Cite this

@article{b1452eec3a9a4fc191987f345c672239,
title = "Detection of heterozygotes in maple-syrup-urine disease: Measurements of branched-chain α-ketoacid dehydrogenase and its components in cell cultures",
abstract = "To detect heterozygotes for maple-syrup-urine disease (MSUD), activities of branched-chain α-ketoacid (BCKA) dehydrogenase and its components in skin fibroblasts of two obligatory heterozygotes and amnion cells of a fetus at risk were measured. Intact heterozygous cells were found to decarboxylate [1-14C]α-ketoisovalerate at rates equal to or only slightly lower than normal subjects. The inability to differentiate heterozygotes from normals with the intact cell assay confirms earlier studies with intact leukocytes using [1-14C]leucine as a substrate. By contrast, measurements of BCKA dehydrogenase activity with disrupted cell suspensions showed MSUD heterozygotes with 30{\%}-60{\%} of normal activity. Moreover, biphasic kinetics in heterozygous cells were observed with increasing substrate concentrations. The altered biphasic kinetics probably reflect expression of the normal allele in the early hyperbolic portion of the curve and of the mutant allele in the later secondary rise at high substrate concentrations. Assays of component activities showed concordant E1 decarboxylase deficiency in both heterozygous- and homozygous-affected cells, whereas the E3, dihydrolipoyl dehydrogenase-component, activity was normal. The above results taken together appear to provide an approach to detection of the heterozygote in MSUD.",
author = "Chuang, {D. T.} and Ku, {L. S.} and Kerr, {D. S.} and Cox, {R. P.}",
year = "1982",
language = "English (US)",
volume = "34",
pages = "416--424",
journal = "American Journal of Human Genetics",
issn = "0002-9297",
publisher = "Cell Press",
number = "3",

}

TY - JOUR

T1 - Detection of heterozygotes in maple-syrup-urine disease

T2 - Measurements of branched-chain α-ketoacid dehydrogenase and its components in cell cultures

AU - Chuang, D. T.

AU - Ku, L. S.

AU - Kerr, D. S.

AU - Cox, R. P.

PY - 1982

Y1 - 1982

N2 - To detect heterozygotes for maple-syrup-urine disease (MSUD), activities of branched-chain α-ketoacid (BCKA) dehydrogenase and its components in skin fibroblasts of two obligatory heterozygotes and amnion cells of a fetus at risk were measured. Intact heterozygous cells were found to decarboxylate [1-14C]α-ketoisovalerate at rates equal to or only slightly lower than normal subjects. The inability to differentiate heterozygotes from normals with the intact cell assay confirms earlier studies with intact leukocytes using [1-14C]leucine as a substrate. By contrast, measurements of BCKA dehydrogenase activity with disrupted cell suspensions showed MSUD heterozygotes with 30%-60% of normal activity. Moreover, biphasic kinetics in heterozygous cells were observed with increasing substrate concentrations. The altered biphasic kinetics probably reflect expression of the normal allele in the early hyperbolic portion of the curve and of the mutant allele in the later secondary rise at high substrate concentrations. Assays of component activities showed concordant E1 decarboxylase deficiency in both heterozygous- and homozygous-affected cells, whereas the E3, dihydrolipoyl dehydrogenase-component, activity was normal. The above results taken together appear to provide an approach to detection of the heterozygote in MSUD.

AB - To detect heterozygotes for maple-syrup-urine disease (MSUD), activities of branched-chain α-ketoacid (BCKA) dehydrogenase and its components in skin fibroblasts of two obligatory heterozygotes and amnion cells of a fetus at risk were measured. Intact heterozygous cells were found to decarboxylate [1-14C]α-ketoisovalerate at rates equal to or only slightly lower than normal subjects. The inability to differentiate heterozygotes from normals with the intact cell assay confirms earlier studies with intact leukocytes using [1-14C]leucine as a substrate. By contrast, measurements of BCKA dehydrogenase activity with disrupted cell suspensions showed MSUD heterozygotes with 30%-60% of normal activity. Moreover, biphasic kinetics in heterozygous cells were observed with increasing substrate concentrations. The altered biphasic kinetics probably reflect expression of the normal allele in the early hyperbolic portion of the curve and of the mutant allele in the later secondary rise at high substrate concentrations. Assays of component activities showed concordant E1 decarboxylase deficiency in both heterozygous- and homozygous-affected cells, whereas the E3, dihydrolipoyl dehydrogenase-component, activity was normal. The above results taken together appear to provide an approach to detection of the heterozygote in MSUD.

UR - http://www.scopus.com/inward/record.url?scp=0020030002&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0020030002&partnerID=8YFLogxK

M3 - Article

C2 - 7081220

AN - SCOPUS:0020030002

VL - 34

SP - 416

EP - 424

JO - American Journal of Human Genetics

JF - American Journal of Human Genetics

SN - 0002-9297

IS - 3

ER -