IN-VITRO MODELING OF THE TERNARY INTERACTION IN JUVENILE-HORMONE METABOLISM

The gradual decline in juvenile hormone (JH) titer followed by its com plete clearance early in the last larval instar is required for the on set of the metamorphosis of lepidopterous larvae. JH titer is regulate d by both biosynthesis and degradation. Two major pathways for JH meta bolism, ester hydrolysis and epoxide hydration, are due to JH esterase (JHE) and JH epoxide hydrolase (JHEH), respectively. In vitro experim ents designed to elucidate the molecular mechanism of JH metabolism ar e described. First, microsomal JHEH in Manduca sexta eggs was identifi ed by using photoaffinity analogs of JH, and purified to homogeneity w ith ion exchange and hydroxylapatite columns. Purified JHEH from M. se xta eggs was kinetically characterized. The effects of pH and various reagents imply that JHEH in M. sexta eggs and mammalian microsomal EH strongly resemble each other. JH binding protein (JHBP) appears to pro tect JH from JHEH; however, the hydration of JH-acid is not affected b y JHBP because JH-acid is not bound by JHBP, but is a good substrate f or JHEH. Thus, the major function of JHEH in M. sexta eggs, which has a cytosolic JHBP, is likely to act as the ultimate scavenger for JH by hydrating JH-acid. Second, although virtually all JH exists as the JH BP JH complex in hemolymph, JHE nonetheless effectively hydrolyzes JH in a JHBP JH complex. Circular dichroism experiments suggest the possi bility of a direct interaction between JHBP and JHE. These experimenta l approaches using reconstituted in vitro model systems may elucidate some of the complex interactions in the JH signaling and metabolic pat hways. (C) 1996 Wiley-Liss, Inc.