Production, optimization, and purification of alkaline thermotolerant protease from newly isolated Phalaris minor seeds

The present work aims to purify and perform a preliminary analysis on a thermostable serine alkaline protease from a recently identified P. minor. The enzyme was purified 2.7-fold with a 12.4 % recovery using Sephadex G-100 chromatography, DEAE-cellulose, and ammonium sulphate precipitation. The isolated enzyme has a specific activity of 473 U/mg. The purified protease had a molecular mass of 29 kDa, and just one band was seen, which matched the band obtained using SDS-PAGE. High thermostability was demonstrated by the enzymes, which had half-lives of 31.79 and 6.0 min (a 5.3-fold improvement), enthalpies of denaturation (Delta H degrees) of 119.53 and 119.35 KJ mol (1), entropies of denaturation (Delta S degrees) of 32.96 and 41.11 J/mol.K, and free energies of denaturation (Delta G degrees) of 108.87 and 105.58 KJ mol(-1) for the protease enzyme. Studies on the folding and stability of alkaline proteases are important since their use in biotechnology requires that they operate in settings of extreme pH and tem-perature. According to the kinetic and thermodynamic properties, the protease produced by P. minor is superior to that produced by other sources and previously described plants, and it might find utility in a variety of in-dustrial fields.

Automated summary

The aim of this study was to purify and analyze a thermostable serine alkaline protease from P. minor. The enzyme was purified 2.7-fold with a 12.4% recovery using chromatography and precipitation techniques. The purified protease showed high thermostability and superior kinetic and thermodynamic properties compared to other sources and previously described plants.