Enzymatic diagnosis of holocarboxylase synthetase deficiency using apo-carboxyl carrier protein as a substrate

Yoichi Suzuki, Yoko Aoki, Osamu Sakamoto, Xue Li, Shigeaki Miyabayashi, Yasuaki Kazuta, Hiroki Kondo, Kuniaki Narisawa

Research output: Contribution to journalArticlepeer-review

24 Citations (Scopus)

Abstract

We developed a simple and sensitive method for assessing holocarboxylase synthetase (HCS) activity that is based on measuring incorporation of [3H]biotin into apo-carboxyl carrier protein, a subunit of acetyl-CoA carboxylase from E. coli. Kinetic analysis of HCS from normal fibroblasts showed that the K(m) for biotin was 260 ± 94 nmol/l (mean ± S.D.; n = 5). In contrast, the K(m) values of HCS from two cell lines derived from patients with HCS deficiency were 7200 and 3700, clearly distinguishable from the control value. The sensitivity of this assay was so high that we were able to characterize a mutant enzyme whose activity had not been previously detected. Our method is useful for enzymatic diagnosis of HCS deficiency and characterization of HCS.

Original languageEnglish
Pages (from-to)41-52
Number of pages12
JournalClinica Chimica Acta
Volume251
Issue number1
DOIs
Publication statusPublished - 1996 Jul 15

Keywords

  • Carboxyl carrier protein
  • Fibroblasts
  • Holocarboxylase synthetase
  • Multiple carboxylase deficiency

ASJC Scopus subject areas

  • Biochemistry
  • Clinical Biochemistry
  • Biochemistry, medical

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