An endogenous inhibitor of folylpolyglutamate synthetase in Neurospora crassa

Abstract

The met-9 mutant (FGSC 552) of Neurospora crassa was examined for ability to synthesize folylpolyglutamates in vivo and in vitro. After culture in defined media, the concentration of mycelial folates was ca 75 % of that found in the Lindegren A wild type (FGSC 853). The derivatives were principally short-chained folates that supported the growth of Lactobacillus casei without conjugase treatment. The levels of polyglutamyl folates in the mutant were only 6–20% of those present in the wild type under comparable growth conditions. Crude mycelial extracts of met-9 had little folylpolyglutamate synthetase (FPGS) activity but ammonium sulphate fractionation resulted in recoveries in the 150–200% range. A protein fraction precipitating at 30–45% of saturation with ammonium sulphate contained a FPGS inhibitor that was more potent on preincubation. The inhibitor was present in other polyglutamate-deficient mutants and in the wild type. This factor was non-dialysable, thermolabile and inactivated by urea and trypsin treatment. The inhibitor fraction lacked significant protease and conjugase activities. Although several enzymes were not affected by preincubation with this protein fraction, the FPGS activities of E. coli, bovine liver and pea cotyledons were strongly inhibited.