Isolation of the Astacin-like metalloprotease coding gene (astl) and assessment of its insecticidal activity against Spodoptera littoralis and Sitophilus oryzae

Abstract

Astacin- like metalloprotease (astl) is a multi-domain metallopeptidase that has protease activity against a number of organisms; including fish, frogs, birds and insects. In this present investigation, the full length of astl cDNA was cloned from spider species, Hasarius adansoni. Sequencing of the cloned astl cDNA proved that its full length including 802 bp with 714bp open reading frame encoding for 238 amino acids. The catalytic domain comprised of 489 nts was cloned and expressed by the yeast expression system Pichia pastoris and its insecticidal activity was determined against two species of agricultural insects Spodoptera littoralis (Lepidoptera:Noctuidae) and Sitophilus oryzae (Coleoptera:Curculionidae). Bioassay was performed using three concentrations (100,500 and 1000 ppm) for four days for S. littoralis and 14 days for S. oryzae. In addition, the astl was fused to the GNA snowdrop lectin in the same frame and expressed in P. pastoris. The synergistic effect of astl and GNA was examined on the S. littoralis larvae and S. oryzae adults. The mortality percentages of the fused protein (Ha-astl/GNA) \"1000 ppm\" after 4 days, were 78.6%{+/-} 4.16 and 71.66% {+/-}3.51 for first and second spodpotera larval instars, respectively. While, lower mortality of the fused protein of the same concentration was observed on S. oryzae adults, 49.3{+/-}2.08 %.