132 HILLEL S. LEVINSON 



phosphatase is near neutrality, setting it off from the cobalt-activated acid 

 pyrophosphatase and the magnesium-activated alkaline pyrophosphatase ex- 

 tracted from Streptococcus faecalis (Oginsky and Rumbaugh, 1955), both 

 from the standpoint of the metal requirement and the pH optimum. 



We have gotten all sorts of relationships in substrate concentration, en- 

 zyme concentration, etc., but these are not really significant now. I should 

 like, though, to indulge in some wild conjectures as to the means by Avhich 

 this enzyme could be involved in spore germination. 



This involves two important reactions. The first reaction is concerned 

 with the synthesis of coenzyme A. and the second is concerned with trans- 

 acetylation. 



(1) ATP -(- Pantoic acid + /3-Alanine ^ Pantothenic acid 



+AMP +PP 



(2) Ad. P-PP + CoASH + Acetate — Ad. P + CoAS-Ac + PP 

 Inorganic pyrophosphate is said by Dr. Fritz Lipmann (1954) to be a 

 product in both reactions. In the presence of pyrophosphatase, the reactions 

 will be forced to the right, due to the disappearance of the pyrophosphate, 

 and the production of CoA and of acetyl CoA will be increased. These 

 increases might result in more rapid utilization of glucose, with increased 

 oxygen uptake. If these reactions can be shown to exist in spores, or in 

 germinating spores, we may have a relationship between the requirements 

 for manganese in spore germination and in pyrophosphatase activation. 

 The possibility exists, and this is totally unsupported, that manganese acts 

 as an activator of proteolytic enzymes resulting in the production of /3-alanine, 

 or of L-alanine convertible to /3-alanine. This would fill a requirement for 

 the production of coenzyme A. In addition, manganese would serve as a 

 pyrophosphatase activator keeping the equilibrium of these reactions to the 

 right. It is well known that the products of an enzvmatic reaction can 

 inhibit the specific enzyme involved in the reaction. Thus, a-amylase is 

 strongly inhibited by a-maltose; fructose and glucose markedly inhibit in- 

 vertase; cellulase is inhibited by cellobiose. Since inorganic phosphate is a 

 product of the enzymatic hydrolysis of pyrophosphate, we might expect 

 phosphate to inhibit pyrophosphatase. This indicates where we might get 

 a glimmer of understanding of the oft-reported inhibition of germination 

 by inorganic phosphate, i.e. through inhibition of pyrophosphatase. There 

 is a suggestion of the actual involvement of pyrophosphatase in germination 

 (Fig. 12), since after spores have germinated (i.e. become stainable), there 

 is a great reduction in the amount of demonstrable pyrophosphatase. Extract 

 of resting spores produces 1,350 fig of orthophosphate P per mg protein N 

 in 10 minutes. Germinated spore extract produces 220 /xg orthophosphate 

 P in the same time, or about 169^ as much as resting spore extract. It may 



