PEPTIDE-BOND SPECIFICITY OF CALPAIN - PROTEOLYSIS OF HUMAN MYELIN BASIC-PROTEIN

In order to determine the peptide bond specificity of calpain, human myelin basic protein (HMBP) was treated with purified calpain of bovine brain. Upon incubation, HMBP component I (HMBP-I) was degraded into several peptides as demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Component I was more susceptible to degradation than components II and III. HMBP degradation products were separated by high performance liquid chromatography (HPLC) and the cleavage sites in HMBP molecules were determined by peptide sequence analysis and by N- and C-terminal analyses. The major cleavage site was found to be (94)Val-(95)Thr with several minor cleavages at (49)Arg-(50)Gly, (18)Ala-(19)Ser, (23)His-(24)Ala, (27)Gly-(28)phe, (59)Asp- (60)Ser, (70)Gly-(71)Ser, (97)Arg-(98)Thr, (110)Ser-(111)Leu, (145)Asp-(146)Ala, and (156)Leu-(157)Gly. These results indicate that calpain is involved in the limited proteolysis of human myelin basic protein and prolonged incubation causes further digestion of the large peptides. (C) 1991 Wiley-Liss, Inc.