13-06: Simple and rapid screening method coupling in vitro expression and hydrolytic activity of cellulases and related enzymes

Lignocellulosic biomass is an abundant renewable natural resource that can serve as an important raw material for the production of biofuels and other bioproducts. However, the costs associated with enzymatic hydrolysis of lignocellulose limits its use as a source of fermentable sugars. Protein engineering and bioprospecting represent promising approaches to obtain cellulases with improved properties under conditions of practical interest. A major bottleneck in screening cellulases is the development of efficient protein-expression methods and techniques for rapid assay of protein libraries. Conventional cell-based cellulase expression methods are time-consuming and laborious. On the other hand, cell-free protein expression is an attractive alternative because of its flexibility and suitability for high-throughput automation. We have developed a high-throughput expression and screening platform by coupling enzyme expression to biocatalytic activity. This approach was applied to identify novel carbohydrate-active enzymes in metagenomic libraries and improved cellulases in directed evolution experiments. The new method represents a powerful platform for high-throughput screening of cellulases and related enzymes.