Characterization of Proteinases Excreted by Bacillus subtilis Marburg Strain during Sporulation

Abstract

Summary. This study has characterized 3 proteolytic enzymes during sporulation by Bacillus subtilis Marburg strain when grown in nutrient broth. A method of purification is described which permits the separation of 2 different proteinases: one belonging to the metal enzyme group and the other to the serine enzyme group. The third enzyme, probably an esterase, showed a high esterolytic activity, but only low proteolytic activity. Determination of the 3 enzymes in a mixture was accomplished by using specific substrates and inhibitors. They were excreted simultaneously between the end of the growth phase until the appearance of the prespores. During this entire period, 20% of the total proteolytic activity was due to the metal proteinase; 80% of the proteolytic activity and 15% of the esterolytic activity was due to the serine proteinase; 85% of the esterolytic activity was the result of the esterase. These findings will contribute to a more complete phenotypic characterization of those mutants of sporulation that appear to be involved in the production of extracellular hydrolytic enzymes.

An RNA Polymerase Mutation Causing Temperature-Sensitive Sporulation in Bacillus subtilis

“…The accumulation of serine and metal proteolytic activities in the culture supernatant fraction, an early sporulation event (1,7,(13)(14)(15), occurred with a similar time course in mutant and wild-type cells at 350 and 460 (Fig. 3).…”

“…3). As judged by appropriate inhibitor experiments, at any given time 75% of the azocasein activity was due to serine protease and 25% of the activity was due to to metal protease (14). The mutant cells slightly underproduced both enzymes at 35°and 460.…”

Purification and Partial Characterization of Hyaluronate Lyase (EC 4.2.2.1) from Propionibacterium acnes

“…In a survey of enzymes produced by P. acnes and related bacteria, Hoeffler (1 977) showed that 73 yo of P. acnes, 18 yo of P. avidum and 33 yo of P. granulosum strains tested produced hyaluronidase. Puhvel & Reisner (1972) have shown that isolates of P. acnes from both normal skin and the skin of patients with cystic acne produce the enzyme in vitro. The pH optima and temperature stability of the enzymes from the various isolates were described in preparations of crude exocellular products, but no attempt was made to purify or characterize the enzyme.…”

Synthesis of OmpA Protein of Escherichia coli K12 in Bacillus subtilis

“…3 were prepared from cells at the early stationary phase of growth. At that time point the level of OmpA protein was near its maximum (to be shown in detail elsewhere) and the amount of exoproteases, which appear during the stationary phase, was still very small (Millet, 1970;M. Sibakov, personal communication).…”