Synthesis of chitin by particulate preparations from Aspergillus flavus.
Cell-free extracts from Aspergillus flavus catalyzed the synthesis of chitin from UDP-GlcNAc. Most of the activity was associated with membrane-rich fractions whereas no activity was detected in the cell walls. Chitin synthetase was activated by fungal acid proteases; animal and plant proteases destroyed it. Upon incubation at 0 C and 28 C chitin synthetase was inactivated, probably by the action of proteases present in the particulate preparations. Maximal activity was obtained at pH 6.6-7.1 and 15 C. Arrhenius plot showed a biphasic curve with the transition at 7 C. E values were 3300 Kcal/mole above this temperature and 15500 Kcal/mole below it. The enzyme was activated by GlcNAc and required a divalent metal, the most active being Mg++. By plotting v vs UDP-GlcNAc concentration a sigmoidal curve was obtained. Km calculated at high substrate concentrations was 20 mM. Chitin synthetase was competitively inhibited by polyoxin D (Ki 6.5 muM) and (Ki 1.35 mM), the latter giving complex kinetics.
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001231:Aspergillus flavus / Q000201:enzymology / Q000378:metabolism*; D002474:Cell-Free System; D002686:Chitin / Q000096:biosynthesis*; D002687:Chitin Synthase / Q000378:metabolism; D004789:Enzyme Activation; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000494:pharmacology; D008670:Metals / Q000494:pharmacology; D010447:Peptide Hydrolases / Q000494:pharmacology; D013696:Temperature; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism
D000900:Anti-Bacterial Agents; D008670:Metals; D002686:Chitin; D014537:Uridine Diphosphate N-Acetylglucosamine; D002687:Chitin Synthase; D010447:Peptide Hydrolases; D008274:Magnesium