Characterization of the 1,3-beta-glucan Synthase of Aspergillus Fumigatus

J Gen Microbiol. 1993 Dec;139(12):3071-8. doi: 10.1099/00221287-139-12-3071.

Abstract

1,3-beta-Glucan synthase activity has been detected in a membrane fraction extracted from the mycelium of the filamentous fungus Aspergillus fumigatus. The enzyme was solubilized by CHAPS and stabilized by filtration on a Bio-gel P30 column. Highest activity was obtained in the early exponential phase of growth. Four factors--GTP, NaF, sucrose and EDTA--added during the extraction procedure, were essential for optimal 1,3-beta-glucan synthase activity. The soluble enzyme preparation was photolabelled with 5-azido-[32P]UDP-glucose and 5-125IASA-UDP-glucose which bind covalently to the enzyme after UV irradiation. These UDP-glucose substrate analogues were competitive inhibitors of the enzyme with a Ki of 1.42 mM and 0.3 mM for 5-azido-UDP-glucose and 5-ASA-UDP-glucose, respectively (Km for UDP-glucose = 1.9 mM). Potential UDP-glucose-binding polypeptides were identified with molecular masses of 31, 50 and 115 kDa.

MeSH terms

  • Affinity Labels
  • Aspergillus fumigatus / enzymology*
  • Aspergillus fumigatus / growth & development
  • Azides / metabolism
  • Azides / pharmacology
  • Binding, Competitive
  • Cholic Acids
  • Chromatography, Gel
  • Detergents
  • Glucosyltransferases / antagonists & inhibitors
  • Glucosyltransferases / isolation & purification
  • Glucosyltransferases / metabolism*
  • Kinetics
  • Membrane Proteins*
  • Schizosaccharomyces pombe Proteins*
  • Solubility
  • Uridine Diphosphate Glucose / analogs & derivatives
  • Uridine Diphosphate Glucose / metabolism
  • Uridine Diphosphate Glucose / pharmacology

Substances

  • Affinity Labels
  • Azides
  • Cholic Acids
  • Detergents
  • Membrane Proteins
  • Schizosaccharomyces pombe Proteins
  • 5-azidouridine 5'-diphosphoglucose
  • 5-(3-(4-azidosalicylamide)allyluridine)- 5'-diphosphoglucose
  • Glucosyltransferases
  • 1,3-beta-glucan synthase
  • 3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate
  • Uridine Diphosphate Glucose