In-frame cDNA library combined with protein complementation assay identifies ARL11-binding partners

PLoS One. 2012;7(12):e52290. doi: 10.1371/journal.pone.0052290. Epub 2012 Dec 18.

Abstract

The cDNA expression libraries that produce correct proteins are essential in facilitating the identification of protein-protein interactions. The 5'-untranslated regions (UTRs) that are present in the majority of mammalian and non-mammalian genes are predicted to alter the expression of correct proteins from cDNA libraries. We developed a novel cDNA expression library from which 5'-UTRs were removed using a mixture of polymerase chain reaction primers that complement the Kozak sequences we refer to as an "in-frame cDNA library." We used this library with the protein complementation assay to identify two novel binding partners for ras-related ADP-ribosylation factor-like 11 (ARL11), cellular retinoic acid binding protein 2 (CRABP2), and phosphoglycerate mutase 1 (PGAM1). Thus, the in-frame cDNA library without 5'-UTRs we describe here increases the chance of correctly identifying protein interactions and will have wide applications in both mammalian and non-mammalian detection systems.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • 5' Untranslated Regions
  • ADP-Ribosylation Factors / metabolism*
  • Base Sequence
  • Carrier Proteins / metabolism*
  • Cell Line
  • Gene Library*
  • Humans
  • Molecular Sequence Data
  • Protein Binding
  • Protein Interaction Mapping*
  • Receptors, Retinoic Acid / metabolism

Substances

  • 5' Untranslated Regions
  • Carrier Proteins
  • Receptors, Retinoic Acid
  • retinoic acid binding protein II, cellular
  • ADP-Ribosylation Factors
  • ARL11 protein, human