Primary structure of a 120 kDa protein associated with the fucose sulfate glycoconjugate constituting the acrosome reaction-inducing substance of thesea urchin, Hemicentrotus pulcherrimus

A fucose sulfate glycoconjugate (FSG), a natural acrosome reaction-inducer, was purified from the egg jelly of the sea urchin Hemicentrotus pulcherrim us. The FSG is composed primarily of four constituents: a 120 kDa protein, a 237 kDa protein, a 258 kDa protein, and a polysaccharide-containing prote in. Among them, the 120 kDa protein was thought to play a critical role in the association of other FSG constituent proteins, and therefore was charac terized from a structural point of view. The protein was isolated from the carboxymethylated FSG by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions, and then digested with trypsin to obtain information regarding the primary structure. Based on the partial amino acid sequences of three internal peptides (FSG120KA: L HNNEYGYGDTAAGEPELAQEEID, FSG120KG: AIDIPAETGHYGR, and FSG120KC: RPTFDLADAVD T) and the N-terminal peptide (LHNNEYGYGDTAAGEPELAQQEID) of the 120 kDa pro tein obtained from intact FSG, degenerate oligonucleotide primers were synt hesized and used to amplify a 297 bp cDNA fragment. This fragment enabled u s to obtain the full-length cDNA (3176 bp) by 5'- and 3'-rapid amplificatio n of cDNA ends. The deduced amino acid sequence revealed that the 120 kDa p rotein is composed of 663 amino acid residues including 72 cysteine residue s, and hence, about 40% is presumed to be carbohydrate by weight. The 120 k Da protein plays an important role in the association of FSG constituent pr oteins (258 and 237 kDa) through disulfide bonds.