Allyl Isothiocynate(AITC)Impact On Sitophilus Zeamais Adult Enzymes And Prediction With COX1 Bingding Sites

Sitophilus zeamais is an important storage pests, a wide range of harm, harm was severe, is the world’s number one storage pests. An extensive range of chemical fumigants, due to resistance or toxicity limitations exist in use, the individual has been banned. Armoracia rusticana(AITC) because of their good insecticidal effect, a small residual amount, minimal damage to theenvironment and the human body, is considered to be has a broad development prospects of plant source fumigants substances.But at the moment about the AITC specific fumigation of Sitophilus zeamais like mechanism is unclear.This study from Sitophilus zeamais adult enzymology, proteomics, gene level for AITC insecticidal mechanisms of fumigation and targets to provide clues. The experiment by measuring the dynamic changes of AITC fumigation Sitophilus zeamais adults four kinds of protease, and the differences in protein after fumigation HCPL- MS and MALDI- TOF/TOF mass spectrometry; Based on RACE and modern molecular simulation technology, this test is designed Sitophilus zeamais cytochrome c oxidase subunit I of the full-length clones and homology modeling and molecular docking with the AITC was, and following results were obtained:1 、 the AITC for Sitophilus zeamais as hydrogen peroxide enzyme, carboxylesterase, mitochondrial respiratory chain complexes II, cytochrome c oxidase activity has obvious dose effect and time effect in the body. Low doses of horseradish can induce the catalase, carboxylesterase, mitochondrial respiratory chain complexes II, the increase of cytochrome c oxidase, while high dose of horseradish, mitochondrial respiratory chain complexes of hydrogen peroxide enzyme II, with cytochrome c oxidase activity; With the increase of processing time, try these two kinds of enzyme activity decreased gradually. 2、after AITC processing Sitophilus zeamais adult,Sitophilus zeamais adult in the changes of protein content, compared with ck, 1μl/L and 2μl/L AITC fumigation processing corn like adult protein content increased significantly; With the increase of corn like adult mortality, corn like Sitophilus zeamais adult body protein content gradually reduced. The SDS-polyacrylamide gel electrophoresis line protein separation and purification, and the differences in protein HPLC- MS and MALDI- TOF/TOF, via the database retrieval, is likely to affect the differences of protein for the cytochrome c oxidase subunit I, similarity65%, 54815.1MW, cytochromec oxidase subunit I,(mitochondrion) [Tribolium castaneum]. 3、Using the method of RACE to get over COX1 gene cDNA sequence. The 1500 bp COX1 gene, encoding 499 amino acids. And lepidoptera curculionidae COX1 gene of other length has been registered in the NCBI Blast homology comparison, discovered COX1 unique four dual-core center(heme a3 / CuB) ion binding sites and five low spin heme chemical binding sites coincide. Have a lot of typical characteristics of COX1 protein family, confirmed that the open reading frame coding corn like COX1, named SzeaCOX1. 4、Using homology modeling method, respectively by three-dimensional crystal structure cattle heart cytochrome c oxidase(PDB: 1 v54)got Sitophilus zeamais COX1 three-dimensional structure, molecular docking with AITC. After using Procheck evaluation AITC allyl part in COX1 by amino acid residuesIle-59, Ile-375, Ile-376, Phe-379, Phe-454 and Phe-6 is composed of a hydrophobic cavity bags to form a strong hydrophobic interaction. More importantly, a sulfur atom isothiocyanate propylene carbonate at the other end with the amino acid residues Gly-13 shaped long as 3.2 ? hydrogen bond interactions.