ABSTRACT
A slow growing inl+/- mutant was isolated from an inositol dependent (inl) Neurospora crassa strain. The latter strain produces defective myo-inositol-1-phosphate synthase which has residual activity. Inositol, similarly to that found in wild and inl mutant strains, represses the enzyme production in the inl+/- strain as well. Withdrawing inositol from the medium results in derepression of the enzyme synthesis. Derepression is hindered by cycloheximide. Inl+/- character in the double mutant is brought about by overproduction of the defective myo-inositol-1-phosphate synthase.
Subject(s)
Carbohydrate Epimerases/genetics , Myo-Inositol-1-Phosphate Synthase/genetics , Neurospora crassa/genetics , Neurospora/genetics , Cells, Cultured , Cycloheximide/pharmacology , Immunoelectrophoresis , Myo-Inositol-1-Phosphate Synthase/biosynthesis , Myo-Inositol-1-Phosphate Synthase/immunology , Neurospora crassa/enzymology , Time FactorsABSTRACT
Immunological experiments were performed to demonstrate myo-inositol-1-phosphate synthase (EC 5.5.1.4) and its assumed defective variant in various Neurospora crassa stains. An enzymatically inactive protein fraction was isolated from the inl-mutant by the same procedure as that of the enzyme. It consisted of several components by gel electrophoresis, and produced a positive immune reaction demonstrated by immunodiffusion using immune sera produced against the enzyme. Using immunodisc gel electrophoresis it produced an immunoprecipitate of slightly lower mobility than the enzyme itself. Similarly, positive immune reactions were obtained with the enzyme using immune sera produced against the protein fraction isolated from the inl- mutant. Enzyme activity was demonstrated both in a strain transformed by wild-type DNA and in a spontaneous revertant. The enzymes were subsequently isolated from both strains, and some properties were compared with those of the wild-type enzyme. The specific activities were lower but the Michaelis constants were nearly the same. The immunodisc gel electrophoretic patterns of these enzymes were similar to that of the protein fraction from the inositol requiring mutant.
