Abstract

A-factor is a microbial hormone that can positively control cell differentiation leading to spore formation and secondary metabolite formation in Streptomyces griseus. To identify a protease that is deeply involved in the morphological and physiological differentiation of Streptomyces, the proteases produced by S. griseus IFO 13350 and its A-factor deficient mutant strain, S. griseus HH1, as well as S. griseus HH1 transformed with the afsA gene were studied. In general, S. griseus showed a higher degree of cell growth and protease activity in proportion to its ability to produce a higher amount of A-factor. In particular, the specific activity of the trypsin of S. griseus IFO 13350 was greatly enhanced more than twice compared with that of S. griseus HH1 in the later stage of growth. The specific activity of the metalloprotease of S. griseus HH1 was greatly enhanced more than twice compared with that of S. griseus IFO 13350, and this observation was reversed in the presence of thiostreptone. However, S. griseus HH1 transformed with the afsA gene showed a significantly decreased level of trypsin and metalloprotease activity compared with that of the HH1 strain. There was no significant difference between S. griseus IFO 13350 and HH1 strain in their chymotrypsin and thiol protease activity, yet the level of leu-aminopeptidase activity was 2 times higher in S. griseus HH1 than in strain IFO 13350. S. griseus HH1 harboring afsA showed a similar level of enzyme activity, however, all the three protease activities sharply increased and the thiol protease activity was critically increased at the end of the fermentation. When a serine protease inhibitor, pefabloc SC, and metalloprotease inhibitor, EDTA, were applied to strain IFO 13350 to examine the in vivo effects of the protease inhibitors on the morphological differentiation, the formation of aerial mycelium and spores was delayed by two or three days.

• Keywords: Streptomyces griseus; A-factor; Trypsin; Metalloprotease