Purification and properties of bovine liver holocarboxylase synthetase

Yasushi Chiba, Yoichi Suzuki, Yoko Aoki, Yoshinori Ishida, Kuniaki Narisawa

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31 Citations (Scopus)


Holocarboxylase synthetase was purified 18,000-fold from bovine liver cytosol by a sequence of ammonium sulfate fractionation, alumina Cγ fractionation, DEAE-Sepharose CL-6B, EAH-Sepharose 4B, Sephacryl S-200 HR, Bio-Gel HTP, and Phenyl-Superose HR 5/5 chromatographies. Holocarboxylase synthetase activity was assayed using apopropionyl-CoA carboxylase from a patient with holocarboxylase synthetase deficiency as a substrate. Apopropionyl-CoA carboxylase was easily prepared from cultured lymphoblasts from this patient. Enzyme activity coincided with a 64,000-Da protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Additionally, the molecular mass of the native enzyme was estimated to be 60,000 Da by gel filtration on Sephacryl S-200 HR. These results suggest that purified holocarboxylase synthetase from bovine liver cytosol is a monomeric enzyme. Its Km value for biotin was estimated to be 13 nM.

Original languageEnglish
Pages (from-to)8-14
Number of pages7
JournalArchives of Biochemistry and Biophysics
Issue number1
Publication statusPublished - 1994 Aug


  • Biotin
  • Holocarboxylase synthetase
  • Purification
  • Subunit structure


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