Patent ID: 6322996
Filing Date: 2001-11-27
Classification: A61K,C07K,C12P

Abstract:
A method of modifying a protein comprising reacting a physiologically active protein having a molecular weight of from 5.times.10.sup.3 to 2.times.10.sup.5 and at least one glutamine residue with an amino group donor or an alkylamine-containing polysaccharide having a molecular weight in the range of 5 kDa to 100 kDa in the presence of transglutaminase, thereby modifying said protein by forming an amido linkage between the carboxyamide group of said glutamine residue and the primary amino group of the alkylamine-containing polysaccharide or the amino group donor, wherein the amino group donor is selected from the group consisting of:(I) NH.sub.2 (CH.sub.2).sub.n T(CH.sub.2).sub.m (OCH.sub.2 CH.sub.2).sub.p OR, having a molecular weight of at least 5 kDa,(II) NH.sub.2 (CH.sub.2).sub.n T(CH.sub.2).sub.m (OCH.sub.2 CH.sub.2).sub.p T(CH.sub.2).sub.n NH.sub.2, having a molecular weight of at least 5 kDa,and wherein n is an integer of 1 to 8, m is an integer of 0 to 2, p is an integer of 1 to 400, T is --O--, --C(O)O--, --OC(O)--, --NHCO--, --OCNH--, --NHCONH--, 'OOCNH-- or --HNCOO--, and R is a hydrogen atom, a lower alkyl group having 1 to 5 carbon atoms or a lower acyl group having 2 to 6 carbon atoms, and ##STR2##wherein n is an integer of 1 to 8, q is an integer of 2 to 6, and R is galactose, glucose or N-acetylgalactosamine,wherein said glutamine residue contained in said physiologically active protein forms an amido linkage between the .gamma.-carboxyamide group of said glutamine residue and the primary amino group of dansylcadaverine if 10 .mu.M of said physiologically active protein and monodansylcadaverine in an amount of 100 equivalents per 1 equivalent of said physiologically active protein are kept at 37.degree. C. for 60 minutes in a 100 mM Tris-HCl buffer containing 10 mM CaCl.sub.2 at a pH value of 7.5, in the presence of the said transglutaminase.