Purification and characterization of the carrier protein for juvenile hormone from the hemolymph of the tobacco hornworm Manduca sexta Johannson (Lepidoptera: Sphingidae).

The larval hemolymph of the tobacco hornworm, Manduca sexta, contains a carrier protein that binds specifically and with high affinity the juvenile hormone, an important regulator of insect development. This protein serves to transport the hormone and to protect it from the action of degradative enzymes during early larval stages. Using hemolymph from the last larval stage, we have isolated a pure carrier protein using acetone precipitation, gel filtration, ion exchange chromatography, and preparative isoelectric focusing. Gel filtration, polyacrylamide gel electrophoresis in sodium dodecyl sulfate, and equilibrium ultracentrifugation established that the carrier protein is a single chain polypeptide of approximately 28,000 daltons. The amino acid composition is unexceptional, and no evidence for hexosamine has been obtained. An ion exchange filter disc assay method was used to determine the formation of the complex between the carrier protein and isotopically labeled juvenile hormone. With this technique it was shown that each carrier protein binds one hormone molecule with a dissociation constant of 4.4 +/- 0.2 X 10(-7) M at 0 degrees.