Substantial improvement of protease production using statistical optimization and Downstream approach from the strain Bacillus toyonensis VKB5 sp., isolated from the host Actinidia delicosa

Abstract

Endophytes are microbes that reside inside the plant without affecting the host. A highly active species of extracellular protease production is Bacillus species. This study aims at optimization and partial purification of fibrinolytic protease enzyme production from the isolate Bacillus toyonensis VKB5 sp. isolated from the fruit Actinidia delicosa. Optimization studies on medium components show that the enzyme production can be achieved in alkaline conditions with nutrient sources mannitol, yeast extract, ammonium chloride. The statistical optimization studies using Plackett-Burman and Response surface methodology determines that the interaction of yeast extract with the mannitol and ammonium chloride enhances the enzyme production. Studies on purification stages describe that the protease purified was 7.4 – fold purer that the crude and 68.9% enzyme was recovered comparing the crude. The molecular weight determined was found to be 21.9 kDa using SDS-PAGE. The purity of the protease was analyzed using HPLC. Further studies on the effect of temperature, pH, inhibitors, detergents and metal ions confirm that the enzyme purified can be alkaline serine protease that withstands thermal condition up to 60°C.