2709碱性蛋白酶水解肽谱的动态研究-轻工技术工程专业毕业论文.docxVIP

ABSTRACTProteases ABSTRACT Proteases are enzymes that can hydrolyze protein，which have been widely used in food，medicine and brewing industry．Different kinds of proteases have different levels of site specifity for the selection of cleavage sites．Proteases from microorganism such as alkaline proteinase used in industry,has weak site specifity, and few studies on the peptide mapping during hydrolysis has been reported．Analysis of the proteolytic peptide mapping is important for the understanding of the proteolytic process and can promote the application ofproteases in food industry． There are two methods for peptide mapping analysis according to the different levels of site specifity of protease based on LC—MS．One is sit—specific method，which used in proteomics is mature and reliable．The other is site-unspecific method，SO far the research were lacked about its reliability．In this paper,the peptides from BSA and soybearl protein isolate digested with trypsin were analyzed by LC-QTOF MS／MS， and then identified by both method．The result indicated that the site-unspecified identification method identified 23％and 54％more peptides than the former from the hydrolysates of bovine serum albumin and soybean protein respectively，suggesting that this method could obtain more information of peptide mapping．The high repeatability of identifying the specified peigtides，reaching above 98％，proving that the site-unspecified method also had promising reliability． In this study,2709 alkaline proteinase was purified though salting out、 ion-exchange chromatography and hydrophobic ineraction chromatography,and it WaS purified to electrophoretic purity． The peptides from BSA and soybean protein isolate digested with the purified 2709 alkaline proteinase were analyzed by LC—QTOF MS／MS，and then identified by site-unspecific method．The result shows that the cleavage site of 2709 alkaline proteinase strongly prefer lysine at C-terminal，and alanine，threonine at N-terminal． This paper provided i