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Identification of Motor Protein Cargo by Yeast 2-Hybrid and Affinity Approaches

  • Yuguo Zhang
  • Rong Wang
  • Holly Jefferson
  • Ann O. Sperry
Part of the Methods in Molecular Biology™ book series (MIMB, volume 392)

Abstract

Identification of the molecular composition of the cargo transported by individual kinesin motors is critical to an understanding of both motor function and regulation of the proper intracellular placement of numerous cellular components including proteins, RNA, and organelles. In this chapter, we describe methods to identify the motor tail sequences responsible for cargo binding by expression of green fluorescent protein (GFP)-motor tail fusion proteins in mammalian cells. In addition, we detail two complementary approaches to identify specific proteins associated with these targeting sequences: a yeast 2-hybrid screen and affinity chromatography.

Key Words

GFP yeast 2-hybrid affinity chromatography kinesin motor protein KIFC1 protein-protein interaction 

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Copyright information

© Humana Press Inc., Totowa, NJ 2007

Authors and Affiliations

  • Yuguo Zhang
    • 1
  • Rong Wang
    • 1
  • Holly Jefferson
    • 1
  • Ann O. Sperry
    • 1
  1. 1.Department of Anatomy and Cell Biology, Brody School of MedicineEast Carolina UniversityGreenville

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