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Journal of Inherited Metabolic Disease

, Volume 28, Issue 6, pp 921–930 | Cite as

Kinetic characterization of human hydroxyacid–oxoacid transhydrogenase: Relevance toD-2-hydroxyglutaric and γ-hydroxybutyric acidurias

  • E. A. Struys
  • N. M. Verhoeven
  • H. J. Ten Brink
  • W. V. Wickenhagen
  • K. M. Gibson
  • C. Jakobs
Article

Summary

We investigated the presence of hydroxyacid–oxoacid transhydrogenase (HOT), which catalyses the cofactor-independent conversion of γ-hydroxybutyrate (GHB) to succinic semialdehyde coupled to reduction of 2-ketoglutarate (2-KG) to D-2-hydroxyglutarate (D-2-HG), in human liver extracts employing [2H6]GHB and 2-KG as substrates. We measured incorporation of 2H in D-[2H]2-HG using GC-MS analyses, providing evidence for HOT activity in humans. Kinetic characterization of HOT was undertaken in forward and reverse directions. We employed [2H6]GHB and [2H4]2-KG as cosubstrates in order to develop a HOT activity assay in cultured human fibroblasts derived from patients with D-2-hydroxyglutaric aciduria. HOT activity was quantified in this system by the measurement of D-[2H5]2-HG production. Fibroblasts derived from patients with D-2-hydroxyglutaric aciduria showed normal HOT activities. Our results provide the first demonstration and preliminary kinetic characterization of HOT activity in human tissues.

Keywords

Public Health Internal Medicine Reverse Direction Metabolic Disease Activity Assay 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© SSIEM and Springer 2005

Authors and Affiliations

  • E. A. Struys
    • 1
  • N. M. Verhoeven
    • 1
  • H. J. Ten Brink
    • 1
  • W. V. Wickenhagen
    • 1
  • K. M. Gibson
    • 2
  • C. Jakobs
    • 1
  1. 1.Metabolic Unit, Department of Clinical ChemistryVU University Medical CenterHV AmsterdamThe Netherlands
  2. 2.Molecular and Medical GeneticsOregon Health & Science UniversityPortlandUSA

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