title: Expression of C5a-like biological activities by the fifth component of human complement (C5) upon limited digestion with noncomplement enzymes without release of polypeptide fragments.
abstract: Experimental conditions required for the expression of maximum C5 activation upon limited trypsin hydrolysis were determined to be 0.008 mol of trypsin/mol C5 in a reaction mixture containing 1 mg C5/ml veronal-buffered saline incubated at 37 degrees C for 30 min. Employing these optimal incubation conditions, the primary or preferred site of trypsin hydrolysis of the C5 alpha-chain resulted in the production of C5 alpha 1 (molecular weight, 90,000) and C5 alpha 5 (molecular weight, 25,000) fragments that remained disulfide bonded to the modified C5 molecule (C5'try). Detailed structural-functional analyses clearly indicated the trypsin-mediated conversion of the C5 alpha-chain to C5 alpha 1 and C5 alpha 5 was responsible for the acquisition of neutrophil lysosomal enzyme-releasing and chemotactic activities. Gel filtration column chromatography under physiological ionic strength, pH 7.4, or in the presence of 0.2% SDS further demonstrated that at least 90% of the total recoverable C5a-like biological activity was mediated by the 210,000 molecular weight forms of trypsin-modified C5. Other physiologically relevant, noncomplement protease enzymes (alpha-thrombin, plasmin, and elastase) also activated C5 to express C5a-like reactivities. Analysis of alpha-thrombin-induced, C5 alpha-chain cleavage events by SDS-polyacrylamide slab gel electrophoresis indicated that the mechanism of alpha-thrombin-activation of C5 is similar to that described for trypsin. Reconciliation of this novel mechanism of C5 activation by trypsin with previously published results, and a discussion of the biological significance of noncomplement enzyme-mediated activation of C5 as it might relate to inflammatory processes in vivo, was presented.
mesh_terms: D002634:Chemotaxis, Leukocyte; D002850:Chromatography, Gel; D003182:Complement C5 / Q000378:metabolism*; D015936:Complement C5a; D005341:Fibrinolysin / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008247:Lysosomes / Q000201:enzymology; D009504:Neutrophils / Q000502:physiology; D010196:Pancreatic Elastase / Q000494:pharmacology; D010446:Peptide Fragments / Q000378:metabolism; D010447:Peptide Hydrolases / Q000494:pharmacology*; D013917:Thrombin / Q000494:pharmacology; D014357:Trypsin / Q000494:pharmacology
chemical_list: D003182:Complement C5; D010446:Peptide Fragments; D015936:Complement C5a; D010447:Peptide Hydrolases; D010196:Pancreatic Elastase; D014357:Trypsin; D013917:Thrombin; D005341:Fibrinolysin