γ-Aminobutyric Acid as a Required Germinant for Mutant Spores of Bacillus megaterium

Abstract

Germinated spores of Bacillus megaterium QM B1551 were irradiated with ultraviolet light, and spore-forming survivors were screened for germination requirements. Spore strains which failed to germinate in a variety of defined solutions germinative for spores of the parent strain were obtained. Mutant spores germinated readily in solutions containing yeast extract or one of numerous complex preparations. y-Aminobutyric acid, obtained from yeast extract by column chromatography, was shown to be required for germination by the mutant spores.-y-Aminobutyric acid and L-alanine at final concentrations of I mm each, in solutions of KI (40 mM), equaled the potency of yeast extract (I mg/ml) in the germination of the mutant spores. One of several other amino acids could be substituted, though less effectively, for L-alanine. a-Aminobutyric acid, ,B-aminobutyric acid, falanine, and 5-aminovaleric acid were ineffective substitutes for-y-aminobutyric acid in mutant spore germination.

Metabolism and the triggering of germination of Bacillus megaterium. Concentrations of amino acids, organic acids, adenine nucleotides and nicotinamide nucleotides during germination

“…Strong support for the role of metabolism in triggering germination came from studies on glutamate degradation during germination of B. megaterium QMB 1551 (Foerster, 1972), in which it was shown that 50% of the glutamate pool of the spore was degraded via 4-aminobutyrate within 30s of exposure to germinants. The role of 4-aminobutyrate and the enzyme producing it (glutamate decarboxylase, EC 4.1.1.15) was further emphasized by studies on a mutant requiring 4-aminobutyrate for germination (Foerster, 1971;Foerster & Foerster, 1973).…”

Spore Pool Glutamic Acid as a Metabolite in Germination

“…Residual pool "C-glutamic acid in germinating spores of strain yOG5. Strong presumptive evidence in support of endogenous glutamic acid as a potentially important metabolite in spore germination derives a priori from the fact that mutant spore strains, having a specific requirement for exogenous y-aminobutyric acid, can be obtained (3). A requirement of y-aminobutyric acid by these strains for spore germination may result from their inability to metabolize (decarboxylate) endogenous glutamic acid.…”

“…Preparation of spores. The medium, growth conditions, and the methods used in the preparation and storage of spore suspensions have been described (3). Techniques used in the preparation of "4C-labeled spores are described below.…”

“…Incubation at 37 C was continued until maximum free spores were obtained (24 to 36 hr). Spores were collected, washed, and stored as described (3).…”

“…However, spore glutamate is released as free amino acid when spores are mechanically ruptured or when heated as aqueous suspensions at temperatures exceeding 90 C (12), suggesting that it is not covalently bonded to spore substance, and is, therefore, potentially available as a metabolite. The possible involvement of glutamic acid as a metabolite in spore germination received special significance with the isolation of mutant strains of B. megaterium with a requirement of 'y-aminobutyric acid for spore germination (3). The require-ment of y-aminobutyric acid by these mutant strains could mean that decarboxylation of endogenous glutamic acid may constitute a critical metabolic event in physiological spore germination in the parent strain, but is blocked in the mutant strains.…”