Site-saturation mutagenesis of Glomerella cingulata cutinase gene for enhanced enzyme thermostability

摘要: Cutinase is an important biocatalyst for various industrial applications. This enzyme which has dual functionality comparable to esterases and lipases, efficient in the hydrolysis of soluble esters emulsified triacylglycerols. Naturally-occurring enzymes usually have disadvantages when applied non-natural catalysis due Glomerella cingulata cutinase thermostability. It postulated that by increasing rigidity at certain amino acid positions showing high mobility based on three-dimensional structure G. cutinase, improvement thermostability will be achieved. The N82 was selected its B-factor value determined via B-FITTER program. Megaprimer PCR employed introduce mutations chosen site randomization using NNK degenerate primers. About 300 transformants were screening positive variants. N82_V14 variant observed more thermostable almost 2-fold increase exposed 50°C 1 hr as compared wild-type enzyme. study may provide valuable information regarding thermal stability cutinases denaturation temperatures.Cutinase 2-f...