Proteinase secretion and growth of Clostridium histolyticum

Abstract

YOUNG cultures of B. fluorescens liquefaciens quantitatively secrete a proteinase into the culture medium, as was found by Virtanen & Tarnanen [1931; 1932] and Virtanen & Suolahti [1937]. In previous work on the proteinase of Cl. histolyticum Weil & Kocholaty [1937] reached a similar conclusion. Recently Maschmann [1938, 1] reported on a bacterial proteinase (B. botulinus, B. Welchii) which he considers to be in the "intracellularly acting proteinase of anaerobic bacteria", differing from the proteinase which is secreted by the bacterial oell. In a later publication on Cl. histolyticum, the same author [Maschmann, 1938, 2] concludes that this microorganism produces three distinct proteinases. These are somewhat vaguely differentiated according to their actions on different protein substrates and their behaviour towards cysteine and other activators. Two of these enzymes are believed to be " extracellular " in character, being secreted in the early stages of bacterialgrowth and to be indifferent towards, or even inhibited by, cysteine and other common activators. The third enzyme is considered to be "intercellular", and to differ from the others in its ability to hydrolyse clupein but not gelatin, and to be activated by-SH compounds. Maschmann claims that the activation phenomena observed by us on culture filtrates of Cl. histolyticum are due to this intracellular proteinase, which is freed only by autolysis of the bacterial cell. Our previous work on this microorganism has been extended and the results are still at variance with the conclusions reached by Maschmann. In cultures of Cl. histolyticum, both the bacterial growth and the proteolytic activity (initial and full') of the cell-free bacterial filtrates, reach a maximum value within

Kinetic Characterization of Affinity Chromatography Purified Clostripain

“…Since the first isolation of clostripain by Kocholaty et al (1938) there have been developed several purification methods starting either with the culture filtrate of C/ostridium histolyticum or with a commercially available collagenase preparation containing approximately 1% of clostripain. The latest purification attempts based on ω-aminoalkylagarose columns (Sepharose-C n NH 2 , π = 4... 8) provided specific activities from 10.8 to 11^kat/mg (Gilles et al, 1979;Kula et a/., 1976).…”

“…It has been used especially for the cleavage of Arg-Pro bonds in peptides and proteins (Labouesse and Gros, 1960;Mitchell, 1968;Kocholaty et al, 1938). The protease was shown to be composed of two polypeptide chains, 43 kDa and 12.5 kDa, associated noncovalently (Gilles et al, 1979).…”

Heterologous expression of the clostripain gene from Clostridium histolyticum in Escherichia coli and Bacillus subtilis: maturation of the clostripain precursor is coupled with self-activation

“…Native clostripain recovered from the culture filtrate of Clostridizrm hi.rto&iczlm strains (Kocholaty, 1938) is ' composed of two polypeptide chains, with molecular masses of 41.7 kDa and 15.4 kDa for the heavy and the light chain, respectively (Gilles e t al., 1979). By application of radiolabelled inhibitors, Cysgl of the heavy chain was identified as the catalytic amino acid residue of the active site (Gilles et al, 1983).…”

Increase in Bacteriophage and Gelatinase Concentration in Cultures of Bacillus Megatherium

“…The curves for the rate of formation of phage (23) and of enzymes (24)(25)(26)(27) by cells or from their precursors (28,45) are similar and more or less autocatalytic in character.…”