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Functional Improvement of Food Proteins and Enzymes by Transglutaminase

通过转谷氨酰胺酶改善食品蛋白质和酶的功能

基本信息

批准号：
62560128
负责人：
IKURA Koji
金额：
$ 1.22万
依托单位：
Kyoto University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1987
资助国家：
日本
起止时间：
1987 至 1988
项目状态：
已结题

项目摘要

1. Isolation of transglutaminase from bovine plasma: Plasma Factor X222 is a zymogenic transglutaminase comprising two catalytic subunits (a_2) and two noncatalytic subunits (b_2). An immunoadsorbent column containing monoclonal antibody to the noncatalytic subunit provided a one-step purification procedure that isolated the catalytic subunit of bovine plasma Factor XIII from plasma with a high yield.2. Amino acid sequence of guinea pig liver transglutaminase: The primary structure of guinea pig liver transglutaminase was deduced from its cDNA sequence and compared with that of the catalytic subunit of human plasma Factor XIII. Several regions of strong homology, including the region surrounding the active site cysteine residue, were observed.3. Amino-terminal processing of guinea pig liver transglutaminase: N- and C-terminal peptides were isolate from protease digests of this enzyme and characterized structurally. These results indicated that this enzyme underwent an N-terminal processing, a removal of the initiator methionine and a subsequent acetylation of newly exposed N-terminal alanine residue.4. Bacterial production of animal transglutaminase: We constructed an expression plasmid pKTGl containing a cDNA of guinea pig liver transglutaminase between NcoI and PstI sites of an expression vector pKK233-2 and produced the enzyme in E. coli. The catalytic properties were mostly same between recombinant and natural transglutaminases.

1.从牛血浆中分离转氨酶：血浆因子X222是一种酶原性转氨酶，包含两个催化亚基（a_2）和两个非催化亚基（b_2）。含有非催化亚基单克隆抗体的免疫吸附柱提供了一步纯化程序，可以从血浆中高产率分离牛血浆因子XIII的催化亚基。2.豚鼠肝转氨酶的氨基酸序列：根据其cDNA序列推导出豚鼠肝转氨酶的一级结构，并与人血浆因子XIII的催化亚基的一级结构进行比较。包括活性位点半胱氨酸残基周围区域在内的几个强同源区域被删除.豚鼠肝转氨酶的氨基末端加工：从该酶的蛋白酶底物中分离N-和C-末端肽并进行结构表征。这些结果表明，该酶经历了N-末端加工，起始蛋氨酸的去除和随后的新暴露的N-末端丙氨酸残基的乙酰化.动物转氨酶的细菌生产：我们在表达载体pKK 233 -2的NcoI和Pst I位点之间构建了含有豚鼠肝转氨酶cDNA的表达质粒pKTG 1，并在E.杆菌重组和天然转氨酶的催化性质基本相同。