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

Juvenile hormone esterase (JHE), which catalyzes the hydrolysis of juvenile hormone, was isolated from the hemolymph of 5(th) instars of Lymantria dis par by two different procedures. One procedure was based on affinity chroma tography and the other on anion-exchange chromatography. The material from both purifications showed bands of approximately 50 kDa when analyzed by SD S-PAGE. Isoelectric focusing (IEF) gels in combination with enzyme activity assays indicated two isoelectric forms with the same pi values (pH 5.1. an d 5.3) from affinity purification and from anion-exchange chromatography. A mino acid sequencing of several internal peptides from the 50 kDa band foll owing affinity purification and alignment of these sequences with JHEs from previously purified lepidopteran species (Heliothis virescens, Manduca sex ta) 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 hemo lymph juvenile hormone esterase from Lymantria dispar. insect Biochemistry 21, 583-595] to hydrolyze JH in the hemolymph of gypsy moth, based on molec ular weight and amino acid sequence. (C) 2000 Elsevier Science Ltd. All rig hts reserved.