Cloning, characterisation, and heterologous expression of the Candida utilis malic enzyme gene

Curr Genet. 2006 Apr;49(4):248-58. doi: 10.1007/s00294-005-0052-z. Epub 2006 Jan 25.

Abstract

The Candida utilis malic enzyme gene, CME1, was isolated from a cDNA library and characterised on a molecular and biochemical level. Sequence analysis revealed an open reading frame of 1,926 bp, encoding a 641 amino acid polypeptide with a predicted molecular weight of approximately 70.2 kDa. The inferred amino acid sequence suggested a cytosolic localisation for the malic enzyme, as well as 37 and 68% homologies with the malic enzymes of Schizosaccharomyces pombe and Saccharomyces cerevisiae, respectively. Expression of the CME1 gene was subject to carbon catabolite repression and substrate induction, similar to the regulatory mechanisms observed for the C. utilis dicarboxylic acid permease. The CME1 gene was successfully expressed in S. cerevisiae under control of the S. cerevisiae PGK1 promoter and terminator. When coexpressed with the S. pombe malate permease gene (mae1), it resulted in a recombinant S. cerevisiae strain able to completely degrade 90% of the extracellular L-malate within 24 h.

MeSH terms

  • Amino Acid Sequence
  • Candida / enzymology*
  • Candida / genetics
  • Cloning, Molecular
  • Fungal Proteins / biosynthesis*
  • Fungal Proteins / genetics
  • Gene Expression*
  • Malate Dehydrogenase / biosynthesis*
  • Malate Dehydrogenase / genetics
  • Molecular Sequence Data
  • Recombinant Proteins / biosynthesis*
  • Recombinant Proteins / genetics
  • Saccharomyces cerevisiae*
  • Schizosaccharomyces
  • Sequence Homology, Amino Acid

Substances

  • Fungal Proteins
  • Recombinant Proteins
  • Malate Dehydrogenase

Associated data

  • GENBANK/DQ173437