Evaluation of proteins involved in germination of toxigenic aspergillus flavus conidia and studies on phytochemicals as anti aflatoxigenic agents

Abstract

Germination is the key event in A. flavus life-cycle, which needs further exploration to develop effective anti-aflatoxigenic compounds and to identify key anti-Aspergillus targets. There are limitations in the A. flavus studies at different morphological conditions at proteomic level. Thus, in our current study toxigenic strainsof A. parasiticus (MTCC8189) andA. flavus (MTCC11866) were treated with four different phytochemicals viz. Gallic acid, Ascorbic acid, Caffeine, Quercetin via Poisoned food plate technique and MTT assay. Results showed antifungal effect of quercetin was the strongest among all in both A. parasiticus (36 µg per ml)andA. flavus (113 µg per ml). Hence, quercetin could be a promising anti-Aspergillus compound. Further, to understand the germination mechanism of A. flavus(7 h) proteomic approach was under takenwhich resulted inproteins from cell wall biosynthesis, metabolisms processes and translation. AF biosynthesis pathway enzymes andMAPK signaling pathway associated proteins were also observed during the germination. Further, to understand quercetin mediated AF biosynthesis inhibition in A. flavus,proteomic as well as HPLC analysis were performed. Thus, A. flavus cultured in corn flour (CF) and A. flavus cultured in corn flour with quercetin (CFQ) revealed expression of 843 and 705 proteins, respectively. cAMP/PKA pathway was expressed in CFQ whereas MAPK pathway in CF. Comparative expression of AF biosynthesizing enzyme revealed the downregulation of PksAin CFQ against CF, which suggested quercetin mediated AF inhibition. HPLC analysis showed decrease in one percent of AFB1 at 7 h to 24 h in CFQ and 51 percent at 48 h. Further, to understand quercetin mediated inhibition of AF biosynthesis at molecular levelhomology modelling and molecular dockingstudies with A. flavusPksA, quercetin and hexanoic acid was undertaken. Binding energy for quercetin was lesser (-7.1kcal per Mol to -5.25kcal per Mol) with respect to hexanoic acid (-4.74 kcal per Mol to -3.54kcal per Mol). Overall results showe