Taewan Kim.

Post-Doctorate Researcher
University of California, Berkeley

Ph.D. Chemical and Biological Engineering, 2007
Seoul National University

Office Location: 220 Calvin Hall

Development of a High-throughput Screening Platform for Cellulase Engineering using a Cell-Free Protein Expression System

Cellulases are important industrial enzymes and have an especial potential for the production of renewable and nonpolluting fuels. However, currently available cellulases have low cellulolytic activity and low thermostability, so that we should produce cellulases with biological properties more suitable for industrial applications.

One of the major challenges in cellulase engineering is the development of an efficient protein expression method as well as a method for the rapid assay of proteins expressed from the variant libraries. Conventional cell-based cellulase expression methods are rather time consuming and labor intensive, and also the efforts to express cellulases in E. coli or yeast have consistently failed to produce active forms.

A cell-free protein expression system, on the other hand, can be used as an alternative protein expression tool to address these problems. This is because the open and flexible nature of the systems allows the addition of other components into reaction mixture depending on the purpose of the experiment. In addition, the PCR-based cell-free protein synthesis enables the high-throughput generation of protein required for cellulolytic activity screening without tedious gene cloning steps, and this technology is readily applicable to automation.

In this work, I am developing a high-throughput cellulase expression and screening platform from the libraries generated by error-prone PCR or DNA shuffling. This platform is able to carry out the overall process including the generation of the PCR product, the functional expression of mutant genes, and the activity test. The proposed method should provide a useful platform for the rapid preparation of protein species for various analyses.