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Molecular and Cellular Biochemistry

, Volume 268, Issue 1–2, pp 9–18 | Cite as

Bovine hexokinase type I: Full-length cDNA sequence and characterisation of the recombinant enzyme

  • Francesca Andreoni
  • Giordano Serafini
  • Maria Elena Laguardia
  • Mauro Magnani
Article
  • 46 Downloads

Abstract

This study reports the revised and full-length cDNA sequence of bovine hexokinase type I obtained from bovine brain. Since dissimilarities have been observed between the published bovine hexokinase type I coding sequence (GenBank accession no. M65140) (Genomics 11: 1014-1024, 1991) and an analysed portion of bovine hexokinase type I gene, the entire open reading frame was re-sequenced and the ends of cDNA isolated by rapid amplification of cDNA ends. The coding sequences, when compared with the published bovine hexokinase type I, contained a large number of mismatches that lead to changes in the resulting amino acid sequence. The revisions result in a hexokinase type I cDNA of 3619 bp that encodes a protein of 917 amino acids highly homologous to human hexokinase type I. The expression of the recombinant full-length enzyme demonstrated that it was a catalytically active hexokinase. When characterised for its kinetic and regulatory properties, it displayed the same affinity for glucose and MgATP as the human hexokinase type I and was inhibited by glucose 6-phosphate competitively versus MgATP. The production of the N- and C-terminal recombinant halves of the enzyme followed by comparison with the full-length hexokinase indicated that the catalytic activity is located in the C-terminal domain. (Mol Cell Biochem 268: 9–18, 2005)

Key words

bovine kinetic properties recombinant hexokinase type I 

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Copyright information

© Kluwer Academic Publishers 2005

Authors and Affiliations

  • Francesca Andreoni
    • 1
  • Giordano Serafini
    • 2
  • Maria Elena Laguardia
    • 2
  • Mauro Magnani
    • 1
    • 2
  1. 1.Centro di BiotecnologieUniversity of UrbinoFanoItaly
  2. 2.Istituto di Chimica Biologica “G. FornainiUniversity of UrbinoUrbinoItaly

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