A Folylpolyglutamate Synthetase Lesion in the met(S2706) Mutant of Neurospora crassa

The met(S2706) mutant (FGSC 4248) of Neurospora crassa was culturedin Vogel's minimal medium with and without L-methionine supplementation.Methionine Stimulated growth but significant mycelial productionalso occurred in minimal medium. The mutant had ability to generatefree and protein methionine from ³⁵S]-cysteine in vivo butthe rate of this synthesis was below that shown by a wild typestrain (Lindegren A, FGSC 853). Cytosolic and mitochondrialfractions of met(S2706) also generated methionine from 3-¹⁴C]-serine.Following dialysis, this reaction was stimulated by additionsof tetrahydrofolate and its diglutamate derivative. Folate analysesshowed that the mutant had a folylpolyglutamate concentrationthat was only 10% of that detected in the wild type. Despitethis, mitochondria of met(S2706) contained folates that werelargely polyglutamates and the total mitochondrial folate concentration(ng/mg protein) was comparable to that of the wild type. Assays of folylpolyglutamate synthetase showed that met(S2706)had a lesion affecting a cytosolic, tetrahydrofolate diglutamate-formingactivity. Cytosolic protein had ability to catalyze a diglutamate hexaglutamate reaction. Mitochondria of the mutant catalyzedthe formation of methylenetetrahydrofolate triglutamate fromthe corresponding methylene monoglutamate. It is suggested thatthe limited folylpolyglutamate synthesis of met(S2706) may involvecytosolic and mitochondrial folylpolyglutamate synthetase activities. (Received September 10, 1984; Accepted December 17, 1984)