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1.发明公开PRODUCTION OF RECOMBINANT PROTEINS BY AUTOPROTEOLYTIC CLEAVAGE OF A FUSION PROTEIN 有权通过融合蛋白的自动切割生产重组蛋白质
公开(公告)号：EP1874932A2
公开(公告)日：2008-01-09
申请号：EP06721223.3
申请日：2006-04-25
摘要： The invention relates to a process for the recombinant production of a heterologous polypeptide of interest, comprising, (i) cultivation of a bacterial host cell which is transformed with an expression vector which comprises a nucleic acid molecule which codes for a fusion polypeptide, the fusion polypeptide comprising a derivative of an autoprotease Npro of Pestivirus, wherein at least one cysteine residue of the naturally occuring autoprotease Npro of Pestivirus is replaced by another amino acid residue, and a second polypeptide which is connected to the first polypeptide at the C-terminus of the first polypeptide in a manner such, that the second polypeptide is capable of being cleaved from the fusion polypeptide by the autoproteolytic activity of the first polypeptide, said second polypeptide being a heterologous polypeptide, wherein cultivation occurs under conditions which cause expression of the fusion polypeptide and formation of corresponding cytoplasmic inclusion bodies, (ii) isolation of the inclusion bodies from the host cell, (iii) solubilization of the isolated inclusion bodies, (iv) induction of autoproteolytic cleavage of the heterologous polypeptide of interest from the fusion polypeptide, and (v) isolation of the cleaved heterologous polypeptide of interest.
摘要翻译： 本发明涉及用于重组生产异源目的多肽的方法，其包括（i）培养用表达载体转化的细菌宿主细胞，所述表达载体包含编码融合多肽的核酸分子，融合物 包含瘟病毒自体蛋白酶Npro的衍生物的多肽，其中瘟病毒的天然存在的自噬蛋白酶Npro的至少一个半胱氨酸残基被另一个氨基酸残基置换，以及第二多肽，其在C末端与第一多肽连接 所述第一多肽以这样的方式存在，即所述第二多肽能够通过所述第一多肽的自体蛋白水解活性从所述融合多肽切割，所述第二多肽是异源多肽，其中培养在引起所述融合多肽表达的条件下发生 和形成相应的细胞质包涵体，（ii） （iii）分离的包涵体的溶解，（iv）诱导来自融合多肽的目的异源多肽的自体蛋白水解切割，和（v）分离目的切割的异源多肽 。
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2.发明授权PRODUCTION OF RECOMBINANT PROTEINS BY AUTOPROTEOLYTIC CLEAVAGE OF A FUSION PROTEIN 有权重组蛋白通过融合蛋白的自我蛋白酶解
公开(公告)号：EP1874932B1
公开(公告)日：2012-10-17
申请号：EP06721223.3
申请日：2006-04-25
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