Application of capillary thin layer isoelectric focusing in polyacrylamide gel to the study of alkaline proteinases in stored-product insects

Abstract

1. 1. Isoelectric focusing in horizontal thin layers of polyacrylamide gel in combination with selected synthetic substrates and inhibitors was used to analyze the alkaline midgut proteinases from larvae of 5 species of stored-product insects. 2. 2. Partially purified proteinases from the black carpet beetle, Attagenus megatoma, had isoelectric points ranging from 3.9 to 9.0. About 15 proteinases were detected when focused gels were incubated on a casein-agar layer. 3. 3. A complex of 17 anionic and cationic proteinases was found in the hide beetle, Dermestes maculatus. Six trypsin-like and two chymotrypsin-like enzymes, one aminopeptidase, and one carboxypeptidase were demonstrated with artificial substrates and inhibitors. 4. 4. Two distinct, strongly anionic proteinases (pI 2.8 and 2.9) were found in Tribolium castaneum, while a single cationic proteinase was demonstrated in the furniture carpet beetle, Anthrenus flavipes. Compared with the other species, there were no strong proteinases in the midguts of larvae of the Indian meal moth, Plodia interpunctella. 5. 5. The use of horizontal isoelectric focusing in polyacrylamide gel for the analysis of proteinases in insects is discussed.