cDNA Libraries pp 211-223

Part of the Methods in Molecular Biology book series (MIMB, volume 729)

Identification of Protein/Target Molecule Interactions Using Yeast Surface-Displayed cDNA Libraries

Protocol

Abstract

We describe a novel expression cloning method based on screening yeast surface-displayed human cDNA libraries by direct affinity interaction to identify cellular proteins binding to a broad spectrum of target molecules. Being a eukaryote, yeast protein expression pathways are similar to those found in mammalian cells, and therefore, mammalian protein fragments displayed on the yeast cell wall are more likely to be properly folded and functional than proteins displayed using prokaryotic systems. Yeast surface-displayed human cDNA libraries have been successfully used to screen for proteins that bind to posttranslationally modified phosphorylated peptides, small signaling molecule phosphatidylinositides, and monoclonal antibodies. In this article, we describe protocols for using yeast surface-displayed cDNA libraries, coupled with fluorescence-activated cell sorting, to select protein fragments with affinity for various target molecules including posttranslationally modified peptides, small signaling molecules, monoclonal phage antibodies, and monoclonal IgG molecules.

Key words

Yeast surface cDNA display Expression cloning Phage antibody Antigen identification Small molecules Posttranslationally modified ligands 

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.UCSF Comprehensive Cancer CenterUniversity of California at San FranciscoSan FranciscoUSA
  2. 2.Department of Anesthesia, UCSF Comprehensive Cancer CenterUniversity of California at San FranciscoSan FranciscoUSA

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