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Purification And Characterization Of Cathepsin B & L From Silver Carp Dorsal Muscle And Research Of Their Proteolytic Action On Myosin

Posted on:2005-06-13Degree:DoctorType:Dissertation
Country:ChinaCandidate:S H LiFull Text:PDF
GTID:1101360122988882Subject:Food Science
Abstract/Summary:PDF Full Text Request
The resources of silver carp {Hypophthalmihthys molitrix) is very rich in our country. So it is necessary to utilizing it. Processing the sliver carp meat into surimi products is of vast range of prospects. However, silver carp is very hard to suwari (setting) and easy to modori (heat-induced softening at 50-70℃), which limited the exploitation and utilization of silver carp resources. According to a great deal of researches, endogenous heat-stable cathepsins B H L in the fish muscle lysosome were implied in the modori phenomenon. So far, whether the endogenous cathepsins in silver carp muscle participated in the heat-induced modori or not is still unknown.Therefore, the rudimental activities of cathepsin B H L in the silver carp mince before and after washing of the meat were studied , and the result showed that the activities of cathepsin B H L were remnant. And both of the remaining activity and remaining percentage of cathepsin L are the highest among three catheptic activities. The autolysis degree of myofibrillar proteins of silver carp dorsal muscle with and without E-64 were investigated respectively, and the results indicated that when without E-64, the changing trend of the relative quantity of myosin heavy chain (MHC) was consistent with that of the rudimentary catheptic activities in the myofibrillar proteins, moreover either E-64 individually or E-64 and PMSF together did not inhibit the autolysis of myosin heavy chain (MHC) thoroughly, suggesting that the modori simulate by the autolysis of myofibrillar proteins at 50℃ might be partly caused by thiol cathepsins in muscle lysosome, as well as by other type of serine proteinases binding to the myofibril probably. Judging from the results above, it comes to the primary conclusion that there was the possibility that cathepsin B, H, L participated in the silver carp surimi softening.In order to illuminate the essential relationship between cathepsin B, L and modori specifically , cathepsin B and cathepsin L were purified and characterized from the silver carp dorsal muscle, and the results indicated that both of the two cathepsins might exist in the form of procathepsin and some processing intermediate, as well as mature monomer of the lowest molecular weight.Adding the same amount (counted according to the unit of activity) of purified cathepsin L and cathepsin B respectively to the silver carp myosin solution, both led to the degradation of MHC at pH6.5 and at 50℃ or 60℃, as well as at 35℃ approximate to the setting temperature in processing. During the reaction time of 0-24h, the degradation of MHC caused by cathepsin L is more evident than that caused by cathepsin B. When the pH value shifted to pH 5.0 and pH 5.5, the proteolytic action of cathepsin L on MHC is much higher than that at pH 6.5, while cathepsin B still hydrolyzed the MHC slightly. All the findings showed that the endogenous lysosomal cathepsin L in silver carp muscle is the more critical proteinase hydrolyzing the surimi protein, and contributes more to the heat-induced softening of silver carp surimi gels. At the same time, the proteolytic action of cathepsin L on MHC might be an agent to difficult-setting at the setting temperature blow 40℃.
Keywords/Search Tags:silver carp, cathepsin L, cathepsin B, purification and characterization, heat-induced softening
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