Purification of a larval settlement-inducing protein complex (SIPC) of the barnacle,Balanus amphitrite

Abstract

Many species of barnacles live gregariously, and chemical cues from conspecifics are believed to induce settlement of cypris larvae. Settlement-inducing factors from the adult barnacle Balanus amphitrite have been partially purified using a nitrocellulose membrane-based assay. The assay was designed for research on active proteins adsorbed to substrata. A settlement-inducing protein complex (SIPC) with a high molecular weight was purified from whole adult barnacles by ammonium sulphate precipitation, ion exchange chromatography on Mono Q, gel filtration on Superdex 200 HR, and lectin affinity chromatography on LCA-Sepharose. SDS-polyacrylamide gel electrophoresis of the purified protein complex showed three major protein bands of 76, 88, and 98 kDa and one minor band of 32 kDa under reducing conditions. Lentil lectin (LCA), a lectin that inhibits adult extract-induced settlement, was found to bind to the subunits of 76, 88, and 98 kDa by lectin blotting. Moreover, we isolated three LCA-binding subunits of SIPC by SDS-PAGE and found that each individual subunit also induced larval settlement. It would appear, therefore, that a specific sugar chain of SIPC plays an important role in the settlement of B. amphitrite. J. Exp. Zool. 281:12–20, 1998. © 1998 Wiley-Liss, Inc.