Photobacterium Producing Collagenases And Their Applications In Preparation Of Collagen Proteins

Collagenase is one kind of enzymes that can degrade native helical collagen fibrils or glutin but not degrade others proteins. Collagenase can cure exceptional hyperplasia of collagen; is used for studies of configuration, biologic synthesize of collagen and preparation of collagen and polypeptide. So the study of collagenase has important academic signification and applied value. Now the means of hot alkali, high pressure, and strong acid have be wash out gradually that are used for producing collagen and polypeptide. Because these means consume time, erode equipment and change the configuration of amino acid. Protease have be used abroad for producing collagen and polypeptide, but trypsin, pepsin and some microbial protease do not incise helix region of native collagen, that the process of producing collagen and polypeptide must have a step that materials of collagen are denaturalized by high temperature. In this way these protease can incise materials of collagen, but the bioactivity of collagen and polypeptide were impacted hugely.Pseudumonas mbl15 was studied that had ability to degrade animal skin. The results of form observe of Pseudumonas mbl15 were short, bacilliform, single or rudder chain;it can move and did not come into being gemma of a fungus. it had flagellum; the long and wide of mbl15 was respectively 1-1.8μm and 0.5-1μm by TEM. The colony was round,orderliness velvet and translucence. The results of physiology and biochemistry Pseudumonas mbl15 showed that mbl15 can hydrolyze amylum and utilize citrate and some glucide, but it's salt endurance was weakly. The optimal fermentation conditions for producing collagenase from Pseudumonas mbl15 were determined by single factor analysis and orthogonal analysis and the results were as follows. The compositions of fermentation medium were:2% beef extract,2% sucrose,0.20% yeast extract, 0.02% CaCl2,0.25% K2HPO4·3H2O,0.05% NaH2PO4·2H2O, The optimal cultivation conditions were 37℃,inoculum 6%,initiate pH 6.5,volume of medium in bottle 10%。Propertise of coarse collagenase from mbl15 protease were studied. The optimal reaction temperature and pH value of the coarse collagenase were 32℃ and 7.5.the activity of collagenase of the coarse collagenase was restrained partially by EDTA和EGTA.. PMSF didn't impact enzyme activity.Fresh pigskin was hydrolyzed applying the coarse collagenase from mbl15. The results showed that. Technology conditions of hydrolyzing pigskin using the coarse collagenase from mbl15 were studied .The optimal conditions of technology conditions were as follows: temperature 35℃, liquor ratio 2.5, hydrolysis time 8h,pH 7.5,mass of enzyme 3%. Character of composition of amino acid and molecular weight of collagen proteins were analysed.