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J Bacteriol. 1968 April; 95(4): 1327-1334
Copyright © 1968 American Society for Microbiology. All Rights Reserved.

Initiation of Bacterial Spore Germination

^a Laboratory of Molecular Biology and Department of Bacteriology, University of Wisconsin, Madison, Wisconsin 53706

ABSTRACT

To investigate the problem of initiation in bacterial spore germination, we isolated, from extracts of dormant spores of Bacillus cereus strain T and B. licheniformis, a protein that initiated spore germination when added to a suspension of heat-activated spores. The optimal conditions for initiatory activity of this protein (the initiator) were 30 C in 0.01 to 0.04 M NaCl and 0.01 M tris(hydroxymethyl)aminomethane (pH 8.5). The initiator was inhibited by phosphate but required two co-factors, L-alanine (1/7 of K_m for L-alanine-inhibited germination) and nicotinamide adenine dinucleotide (1.25 x 10^–4M). In the crude extract, the initiator activity was increased 3.5-fold by heating the extract at 65 C for 10 min, but the partially purified initiator preparation was completely heat-sensitive (65 C for 5 min). Heat stability could be conferred on the purified initiator by adding 10^–3M dipicolinic acid. A fractionation of this protein that excluded L-alanine dehydrogenase and adenosine deaminase from the initiator activity was developed. The molecular weight of the initiator was estimated as 7 x 10⁴. The kinetics of germination in the presence of initiator were examined at various concentrations of L-alanine and nicotinamide adenine dinucleotide.

¹ Present address: Department of Biochemistry, School of Medicine, Stanford University, Palo Alto, Calif. 94303.

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