Preliminary characterization of enzyme activities in Malpighian tubules involved in the breakdown of adipokinetic hormones

Adipokinetic hormones (AKH) from different insect species, crustacean red pigment‐concentrating hormone (RPCH), and synthetic substrates were used to characterized enzyme activities present in the Malpighian tubules (MT) of the desert locuts, Schistocerca gregaria , which are involved in the degradation of AKH. When peptides containing proline (position 6) were incubated with MT homogenate they were cleaved by a post‐proline cleaving enzyme (PPCE). The presence of such an enzyme was confirmed by the breakdown of a synthetic substrate for PPCE. Peptides which do not contain proline were broken down by a post‐phenylalanine cleaving enzyme (PFCE) which could be chymotrypsin or chymotryptic. This PFCE activity(ies) seem(s) to be inactive on the proline‐containig peptides or their fragments or digests these at a slow rate. The C‐terminal chymotrypsin fragments of the AKHs were broken down by MT homogenates with no accumulation of new intermediate products. It is not clear whether another endopeptidase, PPCE, or leucine aminopeptidase (LAP) is responsible. The MTs contain LAP activity; however, this enzyme(s) may be different from its vertebrate counterpart(s). Homogenates of MTs break down equimolar amounts of Pro‐7AMC at approximately the same rate, while porcine kidney LAP (cytosol) cleaved Pro‐7AMC much slower than Leu‐7AMC. The demonstration of carboxypeptidase (CP) A and B activity in the MTs was not possible using conventional substrates such as hippuryl derivatives of amino acids. When CPA from porcine pancreas was added to MT homogenates hippuryl‐phenylalanine was digested proving that the conditions were appropriate for CPA activity to occur. The treatment of a N‐terminally blocked peptide fragment with MT homogenate led to the breakdown of the peptide giving evidence that the MT CP requires a substrate with a somewhat longer length of amino acid residues.