Two alanine aminopeptidase isoenzymes from the developing embryos of the camel tick Hyalomma dromedarii, Koch 1884 (Acari: Ixodidae)

Abstract

ABSTRACTThe specific activity of alanine aminopeptidase was monitored during embryonic development of the camel tick H. dromedarii and a lag period was observed during the first six days. The specific activity was increased gradually until days 15–16 and the maximum level was recorded at age 24–25 days. Two isoenzymes of alanine aminopeptidase were resolved and designated AAP1 and AAP2 and were purified from 24–25 day old eggs of H. dromedarii by chromatography on a DEAE cellulose column and gel filtration on a Sephacryl S-300 column. The purification of AAP1 and AAP2 were increased 9.8-fold and 8.75-fold over the crude extract with recovery percentages of 30.54% and 18.0%, respectively. The molecular weight of both isoenzymes was detected to be 75 ± 2 kDa and the optimum activity of both isoenzymes was found at pH 8. MgCl2 activated AAP1, while MnCl2 and CoCl2 activated AAP2, whereas Bestatin and DTT strongly inhibited AAP1 and AAP2, respectively. Two different isoelectric points were detected at pH 8.2–8.4 and 6.6–6.8 for AAP1 and AAP2, respectively. Both AAP1 and AAP2 isoenzymes did not hydrolyse the synthetic and natural substrates of endopeptidases, indicating the absence of endopeptidase activity. Although both isoenzymes are similar, the activators, inhibitors, and isoelectric point values distinguish them.