COMPLETE AMINO-ACID-SEQUENCE OF CHITINASE-A FROM LEAVES OF POKEWEED (PHYTOLACCA-AMERICANA)

The complete amino acid sequence of pokeweed leaf chitinase-h was dete rmined. First all 11 tryptic peptides from the reduced and S-carboxyme thylated form of the enzyme were sequenced. Then the same form of the enzyme was cleaved with cyanogen bromide, giving three fragments. The fragments were digested with chymotrypsin or Staphylococcus aureus vs protease. Last, the 11 tryptic peptides were put in order. Of seven cy steine residues, six were linked by disulfide bonds (between Cys25 and Cys74, Cys89 and Cys98, and Cys195 and Cys208); Cys176 was free. The enzyme consisted of 208 amino acid residues and had a molecular weight of 22,391. It consisted of only one polypeptide chain without a chiti n-binding domain. The length of the chain was almost the same as that of the catalytic domains of class IL chitinases. These findings sugges ted that this enzyme is a new kind of class IIL chitinase, although it s sequence resembles that of catalytic domains of class IL chitinases more than that of the class IIL chitinases reported so far. Discussion on the involvement of specific tryptophan residue in the active site of PLC-A is also given based on the sequence similarity with rye seed chitinase-c.