STRUCTURAL AND FUNCTIONAL-STUDIES ON BILIVERDIN-ASSOCIATED CYANOPROTEIN FROM THE BEAN BUG, RIPTORTUS-CLAVATUS

Structural and functional analysis were performed on biliverdin-associ ated cyanoprotein (CP) from the hemolymph (CP-1 to CP-4) and eggs (CPe gg, identical to CP-1) of the bean bug, Riptortus clavatus. Isoelectri c focusing analysis of purified CPegg and CP-4 revealed that they are composed of a single alpha subunit and a more acidic beta subunit resp ectively. N-terminal amino acid sequencing revealed that there are six different amino acid residues between the two subunits up to the 29 c ycles determined, indicating that they are encoded by different genes. Appreciable sequence similarities in N-terminal region are found betw een the CP subunits and several insect hexamers. By chemical cross-lin king analysis these CPs were demonstrated to have hexameric structures . Two dimensional peptide mapping revealed that alpha and beta subunit s share common structures in part and that CP-2 and CP-3 are hybrid mo lecules bearing both alpha and beta subunits. From these results the m olecular structure of CPs was established as follows: CP-1 (CPegg)=alp ha(6); CP-2=alpha(4) beta(2); CP-3=alpha(2) beta(4); CP-4=beta(6). In addition, using I-125-labeled CPegg and CP-4, in vivo incorporation in to several tissues was examined during the nymphal-adult development. During the late phase of the 5th instar, CP-4 was sequestered preferen tially over CPegg by the fat body, while after emergence both proteins were suggested to be incorporated into the newly-formed cuticle. CPeg g was sequestered massively by the developing ovaries whereas no appre ciable incorporation of CP-4 was observed.