Collagenolytic Activity of Some Marine Bacteria1

Abstract

Reconstituted, acid-extracted collagen was used to prepare a medium to screen proteolytic marine bacteria for their ability to elaborate collagenolytic enzymes. The medium was resistant to solubilization by trypsin, hyaluronidase, chondroitinase ABC, and various marine proteinases, but was readily hydrolyzed by commercial Clostridium collagenases. Eighty-seven marine isolates collected in the vicinity of Bermuda, Oahu (Hawaii), and Stone Harbor and Cape May, N.J., were screened. Approximately 44% of the isolates were capable of elaborating enzymes that hydrolyzed reconstituted collagen gels. Several cultures produced collagenolytic enzymes only when grown in the presence of collagen or degradation products of collagen, and with very few exceptions the presence of collagen in the medium greatly enhanced collagenolytic enzyme production. The enzymes from a collagenolytic Bermuda marine isolate were studied in more detail to illustrate that the enzymes capable of hydrolyzing reconstituted collagen were separable from nonspecific proteinases by zone electrophoresis and that these enzymes were true collagenases by virtue of their ability to hydrolyze native bovine Achilles' tendon obtained from three different sources.

Induction of collagenase production in Vibrio B-30

“…This medium is referred to as HD½hSW. Other details of culture maintenance and storage were described previously (14).…”

“…Assay of collagenase activity. Acid-extracted, dialyzed calfskin collagen (10 to 13 mg of protein per ml as determined by the method of Lowry et al [10] or by the biuret method [5]) was prepared by the procedure of Gross and Kirk (7), as described previously (14). Dialysis of the extract against cold (0 to 4°C) Sorensen phosphate buffer (pH 7.6; ,u = 0.4) was continued until the pH of the extract was between 7.0 and 7.6.…”

“…Studies in our laboratory concentrated on the isolation of marine organisms capable of producing collagenases. In the course of this work, it was shown that approximately 44% of the marine isolates were capable of elaborating enzymes which hydrolyzed reconstituted collagen gels (14). In this search for collagenolytic marine bacteria, the existence of apparently inducible collagenases was observed.…”

Edema and hemoconcentration in mice experimentally infected with Vibrio vulnificus

“…Many marine Vibrio species produce inducible coilagenases (11), and such enzymes were considered as possible candidates for an edemaproducing toxin. Various nutritional manipulations were suggested by this possibility, as well as by the conditions found optimal (9) for enterotoxin production in Escherichia coli.…”

Improved Tenderness of Restructured Beef Steaks by a Microbial Collagenase Derived from Vibrio B‐30

“…Two custom batches of Vihrio B-30 collagcnase (Merkel et al, 1975) were prcparcd according to the method described by Merkcl and Dreisbach (1978). Briefly, bacteriological medium containing undenatured calf-skin collagen was used to stimulate collagcnasc production by Vihrio B-30 (ATCC, 21250).…”