The Evaluation Method Of Subtilisin’s Thermostability Transformation Effect

The study of protease thermostability modification is becoming a researchhotspot in recent years, which is of great theoretical significance and practical value.On one hand, the work on protease thermostability modification can help peopleexplore the protein thermal stability mechanism more deeply and understand theprotein structure and function more comprehensively. On the other hand, due to thenatural protease has a low tolerance for temperature, which is a serious constraint toprotease’s application in industrial production, making its advantages of high catalyticefficiency and stereoselectivity cannot get played well, so there is an urgent need forbetter thermostability modification methods to improve protease’s thermal stability,and promote the practical application of protease in real life.At present, the experimental method of measuring the heating is mainly used inevaluating the effect of the thermostability modification. This method can determinewhether the thermal stability of protease is improved very well, but it istime-consuming. So there is a need to find a simpler and more efficient way todetermine the effect of protein thermophilic transformation, which can promote thejob of thermostability modification of protease.This article aims to establish a method of evaluating the effect of thethermophilic mutation results. We chose subtilisin BPN’as research object and picked6point mutations of it as research samples. By using molecular dynamics simulationmethod in4different simulated conditions, we compared and analysed someparameters between the wild and6mutated proteins, collected5effective parameterswhich are ΔBOND, ΔELECT, ΔANGLE, ΔVDW and ΔDIHED and defined the firstfour of them as character effective parameters, established the evaluation method ofsubtilisin BPN’s thermostability transformation effect. We used this method onevaluating the subtilisin BPN’s other6mutations and the tryptophan synthasealpha-subunit’s thermostability transformation effects, found the results werecompletely coincided with the existing data, and showed that the method can be usedto evaluate the protease’s thermostability transformation effect. Finally we appliedthis method to the practical work, evaluated the thermostability modifying effects of5random mutants, the results defined all of them were valid mutants.This evaluation method of thermostability transformation by using moleculardynamics simulation, which can evaluate the effects better, more conveniently. It canprovide a theoretical guidance in the future study of protease mutations, and it hashigh significance in protein thermostability transformation studies.