Directed evolution of cellulase from Trichoderma reesei for higher activity and development of microtiter plate assay based on cellobiose dehydrogenase

Abstract

Cellulase (EC 3.2.1.4) are important enzymes in food, paper, textile, detergent and biofuel industries. Most cellulases have low activity and stability. Improving these properties would have substantial impact on numerous industrial processes. Enzymatic properties can be improved by directed evolution, but the screening process is the limiting step. Coupled cellulase assay has been developed in order to improve the screening process. This method does not require boiling samples and allows rapid screening of mutants in a microtiter plate. The aim of this study was to establish enzyme coupled assay where cellulase first hydrolyzes carboxymethylcellulose (CMC), and cellobioses dehydrogenase (CBDH) and dichlorophenolindophenol (DCPIP) is used subsequently for detection of reducing ends