Understanding Phytase Thermostability Bioprospecting and Modulation

Abstract

Phytase, which is myoinositol hexakisphosphate phosphohydrolases, is the enzyme that hydrolyzes phytate (substrate) to release lower myoinositol, micronutrients, and inorganic phosphorous. Phytate exhibits anti-nutritional activity and has the potential to cause environmental issues. Hence, phytase has wide applications in the feed and food industry, aquaculture, myo-inositol phosphate production, bioethanol production, and the soil amendment process. The industrial and soil amendment applications of phytases demand highly stable phytases. Commercial phytase application is also limited due to patent protection and expensive downstream processing. Hence, there is a continuous demand to search for novel stable phytases with catalytic activity to hydrolyze phytate. The present study focused on the bioprospecting of novel phytases via different methods and engineering them to improve their characteristics via a protein engineering approach. Three different bioprospecting methods i.e., conventional, metagenomics, and in-silico were employed to isolate and screen novel phytase producers from culturable, non-culturable, and database sources, respectively. Out of the six isolates obtained via conventional approach, Klebsiella variicola PSD soil isolate was found to belong to the Histidine Acid Phytase. The metagenomics results highlighted the Xanthomonadaceae family as a potential phytase producer. A total of 67 different candidates including Pantoea vagans (PV) and Edwardsiella tarda (ET) were identified as potential phytase producers via In-silico bioprospecting approach. Two potential phytases, P.vagans phytase (PVP) and E.tarda phytase (ETP) from a bioprospecting study were then subjected to in-silico characterization and experimental validation. The results indicated that PVP exhibited greater stability than ETP; hence, it was selected for further optimization using rational engineering techniques.