The degradation of p-nitrophenol (PNP) by Moraxella and Pseudomonas spp. involves an initial monooxygenase-catalyzed removal of the nitro group. The resultant hydroquinone is subject to ring fission catalyzed by a dioxygenase enzyme. We have isolated a strain of an Arthrobacter sp., JS443, capable of degrading PNP with stoichiometric release of nitrite. During induction of the enzymes required for growth on PNP, 1,2,4benzenetriol was identified as an intermediate by gas chromatography-mass spectroscopy (GC-MS) and radiotracer studies. 1,2,4-Benzenetriol was converted to maleylacetic acid, which was further degraded by the I-ketoadipate pathway. Conversion of PNP to 1,2,4-benzenetriol is catalyzed by a monooxygenase system in strain JS443 through the formation of 4-nitrocatechol, 4-nitroresorcinol, or both. Our results clearly indicate the existence of an alternative pathway for the biodegradation of PNP.
A true collagenase was isolated from the culture fluid of a marine bacterium which has been designated Vibrio B-30 (ATCC 21250). Collagenase production was obtained only in media containing collagen or certain degradation products of collagen. Partial purification on DEAE-cellulose and Sephadex G-200 columns produced active enzyme which was free of nonspecific proteases but which contained two collagenases. The two collagenases have the same apparent molecular size, and evidence is presented to support the theory that one collagenase is derived from the other. Vibrio B-30 collagenase appears to be a tetramer with a molecular weight of about 105 000 composed of two different subunits (mol wt 24 000 and 28 000). Some of the properties of the Vibrio collagenase are compared with those of Clostridium histolyticum collagenase. Molecular weights, subunit structures, specificity and mode of collagen hydrolysis, insensitivity to diisopropyl fluorophosphate and calf serum, and sensitivity to certain metal ion complexing agents and isopropyl alcohol are similar for the collagenases from both organisms. However, Vibrio B-30 collagenase and Clostridium collagenase differ immunologically and electrophoretically.
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.
Fascinating insights into how chemistry controls our response to external stimuli using an encounter between a man and a snake to show the macroscopic effects of the molecular reactions. Along the way a lot of organic, structural, and physical chemistry is introduced.
The inducible nature of an extracellular collagenase produced by a marine Vibrio (Vibrio B-30, ATCC 21250) was demonstrated by observing the increase in extracellular collagenase activity after the addition of collagen to cell cultures in the latter part of the exponential growth phase. When collagenase-hydrolyzed collagen was added, the lag time required before collagenase production was detected decreased significantly compared with cultures receiving collagen. Cells
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