Isolation and characterization of juvenile hormone esterase from hemolymph of Lymantria dispar by affinity- and by anion-exchange chromatography

Abstract

Juvenile hormone esterase (JHE), which catalyzes the hydrolysis of juvenile hormone, was isolated from the hemolymph of 5 th instars of Lymantria dispar by two different procedures. One procedure was based on affinity chromatography and the other on anion-exchange chromatography. The material from both purifications showed bands of approximately 50 kDa when analyzed by SDS-PAGE. Isoelectric focusing (IEF) gels in combination with enzyme activity assays indicated two isoelectric forms with the same pI values (pH 5.1. and 5.3) from affinity purification and from anion-exchange chromatography. Amino acid sequencing of several internal peptides from the 50 kDa band following affinity purification and alignment of these sequences with JHEs from previously purified lepidopteran species ( Heliothis virescens, Manduca sexta) showed high homology of these enzymes. The isolated JHE, at least in the stage of insect used, was different from the enzyme reported earlier [Valaitis, A.P., 1991. Characterization of hemolymph juvenile hormone esterase from Lymantria dispar. Insect Biochemistry 21, 583–595] to hydrolyze JH in the hemolymph of gypsy moth, based on molecular weight and amino acid sequence.