Resolution of proteases in the keratinolytic larvae of the webbing clothes moth

The proteases of the larvae of the webbing clothes moth, Tineola bisselliella, were investigated because of this organism's phylogenetic rank as a member of the lower invertebrates, its unique position as one of the relatively few organisms that can digest keratin and its importance as a serious fabric pest. Both the number and nature of different proteolytic enzymes present were investigated and the various activities partially fractionated by ammonium sulphate precipitation and chromatography on DEAE-cellulose and Sephadex G200 columns. A complex mixture of peptidases and proteinases has been found in extracts of whole larvae and has been shown to be associated with the larval digestive tract. The proteinases include metal-chelator-sensitive proteinases (metalloproteinases) and serine proteinases but no SH-proteinases or acid proteinases. The serine proteinases include both trypsin-like and chymotrypsin-like activities. Four major and three minor anionic trypsin-like enzymes and a single major cationic trypsin-like enzyme have been detected. Only a single anionic chymotrypsin-like enzyme appears to be present. The trypsin-like enzymes are unaffected by the naturally occurring proteinase inhibitors, chicken ovomucoid, soybean trypsin inhibitor and lima bean trypsin inhibitor, while the chymotrypsin-like enzyme is inhibited by soybean trypsin inhibitor only. The enzymes resemble the serine proteinases from microorganisms in their pH stability. The peptidases include both aminopeptidase and carboxypeptidase activities and both are present in multiple forms. Sixteen aminopeptidase bands have been detected and all are present in individual larvae. They are not inhibited completely by reagents specific for any of the common active sites, and have different specificity requirements. Two carboxypeptidases have been detected on acrylamide gels and have been completely separated on DEAE-cellulose. No evidence could be found for the existence of any of these proteases as inactive precursors.