Design, expression, and stability of a diverse protein library based on the human fibronectin type III domain.

ABSTRACT: Protein libraries based on natural scaffolds enable the generation of novel molecular tools and potential therapeutics by directed evolution. Here, we report the design and construction of a high complexity library (30 x 10(13) sequences) based on the 10th fibronectin type III domain of human fibronectin (10FnIII). We examined the bacterial expression characteristics and stability of this library using a green fluorescent protein (GFP)-reporter screen, SDS-PAGE analysis, and chemical denaturation, respectively. The high throughput GFP reporter screen demonstrates that a large fraction of our library expresses significant levels of soluble protein in bacteria. However, SDS-PAGE analysis of expression cultures indicates the ratio of soluble to insoluble protein expressed varies greatly for r

SUBMITTER: Olson CA 

PROVIDER: S-EPMC2203324 | biostudies-literature | 2007 Mar

REPOSITORIES: biostudies-literature