重组群落刺激因子蛋白质

摘要

本发明系有关产制一种可刺激灵长类造血母细胞生长及分化之蛋白质，尤其是群落刺激因子 (CSF) ；本发明一方面提供藉重组DNA技术产制CSF蛋白质之方法，有关含表现该蛋白质基因之载体，有关以该载体转形之微生物及细胞系，及有关如此产制之CSF蛋白质。本发明第二方面提供自天然或重组来源单离及纯化CSF蛋白质之方法，如此纯化之CSF蛋白质其纯度及活性水准皆远超过任何先前报导者。

主权项

1.重组主要粒性细胞一巨噬细胞群落刺激因子(GM-CSF)蛋白质。2.以骨髓分析之比活性为至少1 X10 单位/毫克之GM-CSF蛋白质。3.根据申请专利范围第1或2项之GM-CSF蛋白质,其为人之GM-CSF。4.根据申请专利范围第1或2项之GM-CSF蛋白质,其具有如下所示CSF-Thr之胺基酸序列 :MET TRP LEU GLN SER LEU LEU LEU LEUGLY THR VAL ALA CYS THR ILE SERALA PRO ALA ARG SER PRO SER PRO SERTHR GLN PRO TRP GLU HIS VAL ASNALA ILE GLN GLU AIA ARG ARG LEU LEUASN LEU SER ARG ASP THR ALA ALAGLU MET ASN GLU THR VAL GLU VAL ILESER GLU MER PHE ASP LEU GLN GLUPRO THR CYS LEU GLN THR ARG LEU GLULEU TYR LYS GLN GLY LEU ARG GLYSER LEU THR LYS LEU LYS GLY PRO LEUTHR MET MET ALA SER HlS TYR LYSGLN HIS GYS PRO PRO THR PRO GLU THRSER CYS ALA THR GLN THR ILE THRPHE GLU SER PHE LYS GLU ASN LEU LYSASP PHE LEULEU VAL ILE PRO PHEASP CYS TRP GLU RPO VAL GLN GLU5.根据申请专利范围第1或2项之GM-CSF蛋白质,其具有如下所示CSF-Ile之胺基酸序列 :MET TRP LEU GLN SER LEU LEU LEU LEUGLY THR VAL ALA CYS THR ILE SERALA PRO ALA ARGSER PRO SER PRO SERTHR GLN PRO TRP GLU HIS VAL ASNALA ILE GLN GLU ALA ARG ARG LEU LEUASN LEU SER ARG ASP THR ALA ALAGLU MET ASN GLU THR VAL GLU VAL ILESER GLU MET PHE ASP LEU GLN GLUPRO THR CYS LEU GLN THR ARG LEU GLULEU TYR LYS GLN GLY LEU ARG GLYSER LEU THR LYSLEU LYS GLY PRO LEUTHR MET MET ALA SET HIS TYR LYSGLN HlS GYS PRO PRO THR PRO GLU THRSER CYS ALA THR GLN ILE ILE THRPHE GLU SER PHE LYS GLU ASN LEU LYSASP PHE LEU LEU VAL ILE PRO PHEASP CYS TRP GLU RPO VAL GLN GLU6.根据申请专利范围第1或2项之GM-CSF蛋白质,其系长臂猿GM-CSF。7.根据申请专利范围第6项之GM-CSF蛋白质,其具有如下所示CSF-G之胺基酸序列:MET TRP LEU GLN SER LEU LEU LEU LEUGLY THR VAL ALA CYS THR ILE SERALA PRO SER ARG SER PRO SER PRO SERARG GLN PRO TPR GLU HIS VAL ASNALA ILE GLN GLU ALA ARG ARG LEU LEUASN LEU SER ARG ASP THR ALA ALAGLU ILE ASN GLUTHR VAL GLU VAL VALSER GLU MET PHE ASP LEU GLN GLUPRO THR CYS LEU GLN THR ARG LEu GLULEU TYR LYS GLN GLY LEU ARG GLYSER LEU THR LYS LEU LYS GLY PRO LEUTHR MET MET ALA SER HIS TYR LYSGLN HIS CYS PRO PRO THR PRO GLU THRSER CYC ALA THR GLN ILE ILE THRPHE GLU SER PHE LYSGLU ASN LEU LYSASP PHE LEU LEU VAL THR PRO PHEASp CYs TRP GLU PRO VAL GLN GLY8.根据申请专利范围第1或2项之GM-CSF蛋白质,其含有如下所示之以Ala-Pro-Ala-Arg-起始之CSF-Thr,CSF-Ile之胺基酸序列,或以Ala-Pro-Ser-Arg-起始之CSF-G之胺基酸序列:CSF-THR:ALA PRO ALA ARG SER PRO SER PRO SERTHR GLN PRO TRP GLU HIS VAL ASNALA ILE GLN GLU ALA ARG ARG LEU LEUASN LEU SER ARG ASP THR ALA ALAGLU MET ASN GLU THR VAL GLU VAL ILESER GLU MET PHE ASP LEUGLN GLUPRO THR CYS LEU GLN THR ARG LEU GLULEU TYR LYS GLN GLY LEU ARG GLYSER LEU THR LYS LEU LYS GLY PRO LEUTHR MET MET ALA SER HIS TYR LYSGLN HIS CYS PRO PRO THR PRO GLU THRSER CYS ALA THR GLN THR ILE THRPHE GLU SER PHE LYS GLU ASN LEU LYSASP PHE LEU LEU VAL ILE PROPHEASP CYS TRP GLU PRO VAL GLN GLUCSF — ILE:ALA PRO ALA ARG SER PRO SER PRO SERTHR GLN PRO TRP GLU HIS VAL ASNALA ILE GLN GLU ALA ARG ARG LEU LEUASN LEU SER ARG ASP THR ALA ALAGLU MET ASN GLU THR VAL GLU VAL ILESER GLU MER PHE ASP LEU GLN GLUPRO THR CYS LEU GLNTHR ARG LEU GLULEU TYR LYS GLN GLY LEU ARG GLYSER LEU THR LYS LEU LYS GLY PRO LEUTHR MET MET ALA SER HIS TYR LYSGLN HIS CYS PRO PRO THR PRO GLU THRSER CYS ALA THR GLN ILE ILE THRPHE GLU SER PHE LYS GLU ASN LEU LYSASP PHE LEU LEU VAL ILE PRO PHEASP CYS TRP GLU PRO VALGLN GLUCSF — G:ALA PRO SER ARG SER PRO SER PRO SERARG GLN PRO TRP GLU HIS VAL ASNALA ILE GLN GLU ALA ARG ARG LEU LEUASN LEU SERARG ASP THR ALA ALAGLU ILE ASN GLU THR VAL GLU VAL VALSER GLU MET PHE ASP LEU GLN GLUPRO THR CYS LEU GLN THR ARG LEU GLULEU TUR LYS GLN GLY LEU ARG GLYSER LEU THR LYS LEU LYS GLY PRO LEUTHR MET MET ALA SER HIS TYR LYSGLN HIS CYS PRO PRO THR PRO GLU THRSER CYS ALA THRGLN ILE ILE THRPHE GLU SER PHE LYS GLU ASN LEU LYSASP PHE LEU LEU VAL THR PRO PHEASP CYS TRP GLU PRO VAL GLN GLY9.根据申请专利范围第8项之GM-CSF蛋白质,其中以Ala-Pro-Ala-Arg-或Ala-Pro-Ser-Arg-起始之胺基酸序列系以甲硫胺酸残基前导。10. 根据申请专利范围第1或2项之GM-CSF蛋白质,此蛋白质为胺基酸序列对应于天然GM-CSF蛋白质之GM-CSF蛋白质,唯加入、取代或移除一或多个未实质影响天然CM-CSF之生物活性之胺基酸。11. 根据申请专利范围第9项之GM-CSF蛋白质,此蛋白质为具有天然GM-CSF之胺基酸序列之GM-CSF蛋白质,唯系以甲硫胺酸残基前导。