Biosynthesis of UDP-GlcA, a Key Metabolite for Capsular Polysaccharide Synthesis in the Pathogenic Fungus Cryptococcus Neoformans

Biochem J. 2004 Jul 1;381(Pt 1):131-6. doi: 10.1042/BJ20031075.

Abstract

UDP-glucose dehydrogenase catalyses the conversion of UDP-glucose into UDP-GlcA, a critical precursor for glycan synthesis across evolution. We have cloned the gene encoding this important enzyme from the opportunistic pathogen Cryptococcus neoformans. In this fungus, UDP-GlcA is required for the synthesis of capsule polysaccharides, which in turn are essential for virulence. The gene was expressed in Escherichia coli and the 51.3-kDa recombinant protein from wild-type and five mutants was purified for analysis. The cryptococcal enzyme is strongly inhibited by UDP-xylose and NADH, has highest activity at pH 7.5 and demonstrates Km (app) values of 0.1 and 1.5 mM for NAD+ and UDP-glucose respectively. Its activity was significantly decreased by mutations in the putative sites of NAD+ and UDP-glucose binding. Unlike previously reported eukaryotic UDP-glucose dehydrogenases, which are hexamers, the cryptococcal enzyme is a dimer.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis Proteins / chemistry
  • Bacterial Proteins / chemistry
  • Cloning, Molecular / methods
  • Conserved Sequence / genetics
  • Cryptococcus neoformans / enzymology*
  • Cryptococcus neoformans / pathogenicity
  • DNA, Complementary / genetics
  • DNA, Fungal / genetics
  • Fungal Proteins / biosynthesis
  • Fungal Proteins / chemistry
  • Fungal Proteins / genetics
  • Humans
  • Molecular Sequence Data
  • Polysaccharides / biosynthesis*
  • Polysaccharides / genetics
  • Sequence Alignment / methods
  • Uridine Diphosphate Glucose Dehydrogenase / chemistry
  • Uridine Diphosphate Glucose Dehydrogenase / genetics
  • Uridine Diphosphate Glucose Dehydrogenase / metabolism*
  • Uridine Diphosphate Glucuronic Acid / biosynthesis*

Substances

  • Arabidopsis Proteins
  • Bacterial Proteins
  • DNA, Complementary
  • DNA, Fungal
  • Fungal Proteins
  • Polysaccharides
  • Uridine Diphosphate Glucuronic Acid
  • Uridine Diphosphate Glucose Dehydrogenase

Associated data

  • GENBANK/AAK95561