Initiation of Bacterial Spore Germination

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 ( p H 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 × 10 −4 m ). 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 −3 m 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 × 10 4 . The kinetics of germination in the presence of initiator were examined at various concentrations of l -alanine and nicotinamide adenine dinucleotide.