Characterization of the solubilized oocyte membrane receptor for insecticyanin, a biliprotein of the hawkmoth, Manduca sexta.
We report the solubilization and characterization of the oocyte membrane receptor for insecticyanin, a blue biliprotein of the hawkmoth Manduca sexta. The insecticyanin receptor was solubilized using 40 mM CHAPS. Strong binding affinity of [125I]insecticyanin to its solubilized receptor was demonstrated to be heat-labile, pH-dependent, Ca2+-dependent, and saturable. The binding was inhibited by excess unlabeled insecticyanin, but not by two other major hemolymph and oocyte proteins, vitellogenin and lipophorin. The receptor for insecticyanin showed tissue specificity: it was present only in oocyte membranes, not in membranes of fat body, midgut or ovariole sheath. The equilibrium data for the solubilized receptor, K(d) and B(max), were estimated to be 17 nM and 11.4 pmol/mg solubilized proteins, respectively. The results from co-immunoprecipitation showed that the apparent molecular mass for the insecticyanin receptor is approximately 185 kDa while chemical crosslinking of the insecticyanin-receptor complex revealed a product with a molecular mass near 10(3) kDa. This suggests that the insecticyanin receptor has a multimeric structure, or that four receptor molecules can bind to one insecticyanin tetramer.
Kang
Y
Y
Department of Biochemistry, University of Arizona, Tucson 85721, USA.
Ziegler
R
R
van Antwerpen
R
R
Law
J H
JH
D000818:Animals; D002793:Cholic Acids; D003432:Cross-Linking Reagents; D003902:Detergents; D004527:Egg Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D005260:Female; D019476:Insect Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D007447:Invertebrate Hormones / Q000378:metabolism*; D018402:Manduca / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D009865:Oocytes / Q000737:chemistry*; D009928:Organ Specificity; D012995:Solubility
D002793:Cholic Acids; D003432:Cross-Linking Reagents; D003902:Detergents; D004527:Egg Proteins; D019476:Insect Proteins; D007447:Invertebrate Hormones; D008565:Membrane Proteins; C043409:insecticyanin protein, insect; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate