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標題: Application potency of engineered G159 mutants on P1 substrate pocket of subtilisin YaB as improved meat tenderizers
作者: Yeh, C.M.
Yang, M.C.
Tsai, Y.C.
關鍵字: application potency;protein-engineered subtilisin YaB;G124;G159;alkalophilic bacillus strain;alkaline elastase;muscle;substitution;specificity;tenderness;gene
Project: Journal of Agricultural and Food Chemistry
期刊/報告no:: Journal of Agricultural and Food Chemistry, Volume 50, Issue 21, Page(s) 6199-6204.
A serine protease, subtilisin YaB, produced by alkalophilic Bacillus YaB, shows promises as a potent meat tenderizer, because its substrate specificity is for small amino acids, which are found at high levels in meat connective tissue proteins. Substrate specificity engineering of the substrate binding pockets was used to generate more suitable meat-tenderizing mutants, G124A, G124V, G159A, and G159S, derived from recombinant wild subtilisin YaB and expressed in Bacillus subtilis DB104. The characteristics of these recombinant enzymes were studied to evaluate their usefulness as improved meat tenderizers. The proteolytic activities of recombinant subtilisin YaB, engineered subtilisin YaBs, and commercially available papain, bromelain, collagenase, and elastase were compared using elastin, collagen, casein, and myofibrillar proteins as substrates. Hydrolysis of beef proteins was evaluated using the myofibrillar fragmentation index and collagen solubility. The results demonstrated that recombinant mutant G159A was the most improved meat tenderizer and can be used in the meat pH range of 5.5-6.0 and the temperature range of 10-50 degreesC. Contrary to the result obtained from artificial substrate, mutant enzymes engineered on G124 residues did not exhibit better tenderizing ability when elastin, collagen, or meat was used as substrate, suggesting the necessity of evaluation by real substrate before protein-engineered enzymes are applied commercially.
ISSN: 0021-8561
DOI: 10.1021/jf0256889
Appears in Collections:食品暨應用生物科技學系

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