Combinatorial library approaches for improving soluble protein expression in Escherichia coli.
High-throughput screening methodologies are already used in structural biology to define efficient protein crystallization and expression conditions. Recently, screening approaches have been extended to the optimization of genetic constructs for improved soluble protein expression. With similarities to the directed evolution strategies used in protein engineering, a target gene encoding a poorly expressed protein is mutated by truncation, fragmentation or point mutation. Rare clones with improved protein expression characteristics are then isolated from the random library using a phenotypic screen or selection. This article reviews the progress in this field and provides a general overview of relevant mutation methods, screens and selections.
Hart
Darren J
DJ
EMBL Grenoble Outstation, 6 Rue Jules Horowitz, 38042 Grenoble, France.
Tarendeau
Franck
F
D000818:Animals; D020650:Combinatorial Chemistry Techniques; D004247:DNA / Q000737:chemistry / Q000235:genetics; D019020:Directed Molecular Evolution / Q000379:methods; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D006801:Humans; D016297:Mutagenesis, Site-Directed; D015202:Protein Engineering / Q000379:methods*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011995:Recombination, Genetic
D011506:Proteins; D004247:DNA