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1. 发明公开

EP1076097A1 METHOD FOR CONTROLLING CLEAVAGE BY OmpT PROTEASE 有权
标题翻译： VERFAHREN ZUR KONTROLLE DER SPALTUNG MIT OMPT PROTEASE
公开(公告)号：EP1076097A1
公开(公告)日：2001-02-14
申请号：EP00906696.0
申请日：2000-03-03
申请人： SUNTORY LIMITED
发明人： OKUNO, Kazuaki , YABUTA, Masayuki , OHSUYE, Kazuhiro
IPC分类号： C12P21/02 , C12N15/09
CPC分类号： C07K14/585 , C07K14/58 , C07K14/605 , C07K2319/50 , C07K2319/75 , C12N15/62 , C12N15/63
摘要： By clarifying the properties of OmpT protease, a novel use thereof as a protease and a novel method of producing a target polypeptide are provided. More particularly, the present invention relates to a method of controlling cleavage of a polypeptide by OmpT protease which comprises converting a sequence site consisting of two arbitrary consecutive amino acids and/or amino acid(s) in the vicinity of said site in said polypeptide into other amino acids, characterized by (1) setting lysine or arginine as the amino acid at the -1-position concerning said site and setting a specific amino acid as the amino acid at the +1-position; and/or (2) setting specific amino acid(s) as the amino acid(s) at the -4-position and/or the -6-position relative to said site; so that desired parts of said polypeptide are cleaved by OmpT protease and/or undesired parts of said polypeptide are not cleaved by ompT protease. For example, the present invention provides a method of treating by OmpT protease a fusion protein having an amino acid sequence wherein the amino acid at the +1-position is X (wherein X is an amino acid other than glutamic acid, aspartic acid or proline); and a method of controlling cleavage by converting the amino acids at the -6-position and the -4-position concerning the cleavage site into amino acids other than acidic amino acids.
摘要翻译：通过澄清OmpT蛋白酶的性质，提供了其作为蛋白酶的新用途和产生靶多肽的新方法。更具体地，本发明涉及通过OmpT蛋白酶控制多肽切割的方法，其包括将所述多肽所述位点附近的两个任意连续的氨基酸和/或氨基酸组成的序列位点转化成其它氨基酸，其特征在于：（1）将赖氨酸或精氨酸设置为与所述位点相关的-1位氨基酸，并将特定氨基酸作为氨基酸位于1位; 和/或（2）将相对于所述位点的-4位和/或-6位上的氨基酸设置为特定氨基酸; 使得所述多肽的所需部分被OmpT蛋白酶切割和/或所述多肽的不需要部分不被ompT蛋白酶切割。例如，本发明提供了通过OmpT蛋白酶处理具有氨基酸序列的融合蛋白的方法，其中+ 1位的氨基酸为X（其中X为除谷氨酸以外的氨基酸，天冬氨酸或脯氨酸）; 以及通过将关于切割位点的-6位和-4位的氨基酸转化成除了酸性氨基酸以外的氨基酸来控制裂解的方法。

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