公开(公告)号：US4782137A

公开(公告)日：1988-11-01

申请号：US76811

申请日：1987-07-23

申请人： Thomas P. Hopp ,  Susan L. Bektesh ,  Paul J. Conlon, III ,  Carl J. March

发明人： Thomas P. Hopp ,  Susan L. Bektesh ,  Paul J. Conlon, III ,  Carl J. March

IPC分类号： C07K14/55 ,  C07K16/44 ,  C12N15/62 ,  A61K37/02 ,  C12P21/00

CPC分类号： C07K14/55 ,  C07K16/44 ,  C12N15/62 ,  C07K2319/00 ,  C07K2319/40 ,  C07K2319/50

摘要： A hybrid polypeptide composed of an identification peptide and a desired functional protein are produced by recombinant DNA techniques. A DNA expression vector is constructed that includes segments of DNA coding for the identification peptide and the desired functional protein. The identification peptide consists of a highly antigenic N-terminal portion and a C-terminal linking portion that connects the identification peptide to the N-terminal of the functional protein. The linking portion of the identification peptide is cleavable at a specific amino acid residue adjacent the functional protein by use of a sequence specific proteolytic enzyme or chemical proteolytic agent. The hybrid polypeptide expressed by the host cells transformed by the cloning vector is removed therefrom and purfied by affinity chromatography techniques by use of an immobilized ligand specific to the antigenic portion of the identification peptide. The protein is then cleaved from the isolated hybrid polypeptide with an appropriate proteolic enzyme or chemical agent, thereby releasing the mature functional protein in highly purified, highly active state.

摘要翻译： 通过重组DNA技术产生由鉴定肽和所需功能性蛋白质组成的杂合多肽。 构建DNA表达载体，其包括编码鉴定肽和所需功能性蛋白质的DNA片段。 识别肽由高度抗原的N末端部分和连接识别肽与功能性蛋白质的N末端的C-末端连接部分组成。 识别肽的连接部分可通过使用序列特异性蛋白水解酶或化学蛋白水解剂在与功能蛋白相邻的特定氨基酸残基处可切割。 由克隆载体转化的宿主细胞表达的杂合多肽从其中除去，并通过使用对鉴定肽的抗原部分特异性的固定化配体的亲和层析技术进行纯化。 然后用合适的蛋白酶或化学试剂从分离的杂交多肽切割蛋白质，从而释放高度纯化，高活性状态的成熟功能性蛋白质。

公开(公告)号：US4703004A

公开(公告)日：1987-10-27

申请号：US573825

申请日：1984-01-24

申请人： Thomas P. Hopp ,  Susan L. Bektesh ,  Paul J. Conlon, III ,  Carl J. March

发明人： Thomas P. Hopp ,  Susan L. Bektesh ,  Paul J. Conlon, III ,  Carl J. March

IPC分类号： G01N33/531 ,  A61K35/74 ,  A61K36/06 ,  A61K38/00 ,  A61K39/00 ,  C07C15/04 ,  C07H21/04 ,  C07K14/005 ,  C07K14/195 ,  C07K14/55 ,  C07K16/00 ,  C07K16/44 ,  C07K19/00 ,  C12N5/00 ,  C12N5/10 ,  C12N15/00 ,  C12N15/09 ,  C12N15/62 ,  C12P21/00 ,  C12P21/02 ,  C12P21/08 ,  C12R1/19 ,  C12R1/865 ,  C12R1/91 ,  G01N33/577 ,  C12N1/00

CPC分类号： C07K14/55 ,  C07K16/44 ,  C12N15/62 ,  C07K2319/00 ,  C07K2319/40 ,  C07K2319/50 ,  Y10S930/28

摘要： A hybrid polypeptide composed of an identification peptide and a desired functional protein are produced by recombinant DNA techniques. A DNA expression vector is constructed that includes segments of DNA coding for the identification peptide and the desired functional protein. The identification peptide consists of a highly antigenic N-terminal portion and a C-terminal linking portion that connects the identification peptide to the N-terminal of the functional protein. The linking portion of the identification peptide is cleavable at a specific amino acid residue adjacent the functional protein by use of a sequence specific proteolytic enzyme or chemical proteolytic agent. The hybrid polypeptide expressed by the host cells transformed by the cloning vector is removed therefrom and purified by affinity chromatography techniques by use of an immobilized ligand specific to the antigenic portion of the identification peptide. The protein is then cleaved from the isolated hybrid polypeptide with an appropriate proteolic enzyme or chemical agent, thereby releasing the mature functional protein in highly purified, highly active state.

摘要翻译： 通过重组DNA技术产生由鉴定肽和所需功能性蛋白质组成的杂合多肽。 构建DNA表达载体，其包括编码鉴定肽和所需功能性蛋白质的DNA片段。 识别肽由高度抗原的N末端部分和连接识别肽与功能性蛋白质的N末端的C-末端连接部分组成。 识别肽的连接部分可通过使用序列特异性蛋白水解酶或化学蛋白水解剂在与功能蛋白相邻的特定氨基酸残基处可切割。 由克隆载体转化的宿主细胞表达的杂合多肽从其中除去，并通过亲和层析技术通过使用对鉴定肽的抗原部分特异性的固定化配体进行纯化。 然后用合适的蛋白酶或化学试剂从分离的杂交多肽切割蛋白质，从而释放高度纯化，高活性状态的成熟功能性蛋白质。