Cloning, expression and characterization of arcelin and its impact on digestive enzymes of the stored product insect pest, Callosobruchus maculatus (F.)

Abstract

Key message The defensive role of arcelin from Phaseolus lunatus for its protection against insect pest Callosobruchus maculatus was elucidated. The potent insecticidal property of arcelin and its impact on pest molecular physiology has identified the adaptive strategies. Abstract The pulse beetle Callosobruchus maculatus causes potential damage to legume crops by infesting the seeds and causes a reduction in the content of total protein. Arcelin found in the wild accessions of the common bean, is an insecticidal protein that has the potency to hamper the metabolism of the bruchid beetle. The arcelin gene from the wild accession of Phaseolus lunatus was isolated and the ORF encoding 158 amino acids was cloned in pET-45b (+) vector. The recombinant was transformed into BL21 STAR (DE3) pLysS cells, and the expressed arcelin was purified using Ni-NTA column. The recombinant protein was used in preparing artificial diet and the insecticidal activity was elucidated against the bruchid pest C. maculatus. Adult emergence and seed damage were drastically reduced in the treated groups. The response towards ingested diet by digested enzymes was elucidated through quantitative gene expression. Maximum expression was observed in the aminopeptidase, followed by upregulation of alpha-amylase, glycoside hydrolase family 31 and cathepsin D-like aspartic protease, and downregulation of cathepsin L-like cysteine protease. These results showed the anti-metabolic nature of the recombinant arcelin. The changes in digestive enzymes to counteract the anti-nutritional nature of the protein were the strategies of the insect defense mechanism.