The amino-acid sequence of purified recombinant pro-major basic protein from Chinese hamster kidney cells was determined to verify the primary structure and glycosylation sites. Reduced and S-carboxamidemethylated protein was first digested with Achromobacter proteinase I. Each peptide was characterized by amino-acid analysis and amino-acid sequence analysis. We could identify all the peptides which were expected from the pro-major basic protein cDNA sequence. Sequence analysis and deglycosylation study revealed that Ser-8, Thr-9, Ser-46 and Asn-70 were glycosylated. The results indicated that proMBP has three types of sugar chains, O-glycoside, N-glycoside and glycosaminoglycan, in the pro-portion.