Limited Proteolysis of the 94000‐Dalton Subunit of Panulirus interruptus Hemocyanin the Carbohydrate Attachment Site

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Abstract
Limited proteolysis of the native monomeric 94‐kDa subunit of Panulirus interruptus hemocyanin by trypsin, plasmin and subtilisin produces an 18‐kDa fragment, a 71‐kDa fragment and a small glycopeptide. In the plasmin digest a 23‐kDa precursor of the 18‐kDa fragment has been observed. Automatic Edman degradations demonstrated that the 18‐kDa fragments have their origin at the N terminus of the 94‐kDa subunit and incubations with carboxypeptidase A showed that the 71‐kDa fragments originate from the C terminus. The glycopeptide is situated in between. The amino acid sequence of the glycopeptide has been determined. Its carbohydrate content accounts for the total carbohydrate of the 94‐kDa subunit.All three proteases cleave the subunit at two positions within a very restricted area of the polypeptide chain, which indicates the presence of an exposed loop, carrying the carbohydrate chain, in the corresponding part of the native molecule.