CHARACTERIZATION OF THE MIDGUT DIGESTIVE PROTEINASE ACTIVITY OF THE 2-SPOT LADYBIRD (ADALIA-BIPUNCTATA L.) AND ITS SENSITIVITY TO PROTEINASE-INHIBITORS

Proteinase activities in the midguts of adult and larval two-spot lady birds were investigated. Both extracts showed a slightly acidic pH opt imum for proteolysis against azocasein (pH 6.0 and pH 5.5 for larvae a nd adults respectively), which correlated well with the physiological pH of the midgut. The proteolytic activity of the larval midguts, dete cted by hydrolysis of Z-phe-arg-pNA, was almost totally inhibited by t he cysteine proteinase inhibitor E-64. Inhibitors diagnostic for the o ther mechanistic classes of proteinase had little or no effect on prot eolysis. In-gel assays showed that the proteolytic activity was due to three major cysteine proteinases with indicated mel. wts. 23, 30 and 55 kDa. In addition to cysteine proteinases, adult ladybirds also poss essed a metalloproteinase activity. Plant protein inhibitors of serine proteases had little effect on ladybird proteolytic activity, but cys tatins (cysteine protease inhibitors) from plant and animal soul ces s howed significant inhibition (up to 90% at 2 mu M). The results sugges t that ladybird digestion may be affected by cysteine proteinase inhib itors expressed in transgenic plants for pest control, via the tritrop hic interaction that occurs between ladybirds, aphids and crop plants. (C) 1998 Elsevier Science Ltd. All rights reserved.