title: Selection of proteins with desired properties from natural proteome libraries using mRNA display.
abstract: mRNA display is a powerful yet challenging in vitro selection technique that can be used to identify proteins with desired properties from both natural proteome and combinatorial polypeptide libraries. The physical conjugation between a protein and its own RNA presents unique challenges in manipulating the displayed proteins at a low nanomolar scale in an RNase-free environment. The following protocol outlines the generation of cDNA libraries derived from natural organisms as well as the steps required for generation of mRNA-protein fusion molecules, in vitro functional selection and regeneration of the selected cDNA library. The selection procedures for the identification of protease substrates and Ca(2+)-dependent calmodulin-binding proteins from natural cDNA libraries are presented as examples. The method can be generally applied to the identification of protein sequences with desired properties from various natural proteome libraries. One round of mRNA display-based selection can be accomplished in ~7 d.
authors.affiliation: Eshelman School of Pharmacy and Carolina Center for Genome Sciences, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
authors.forename: Steven W
authors.initials: SW
authors.lastname: Cotten
authors.forename: Jianwei
authors.initials: J
authors.lastname: Zou
authors.forename: C Alexander
authors.initials: CA
authors.lastname: Valencia
authors.forename: Rihe
authors.initials: R
authors.lastname: Liu
mesh_terms: D020869:Gene Expression Profiling / Q000379:methods*; D015723:Gene Library; D020543:Proteome; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism*
chemical_list: D020543:Proteome; D012333:RNA, Messenger