The phox homology (PX) domain protein interaction network in yeast.
The phox homology (PX) domain is a phosphoinositide-binding domain that is conserved from yeast to human. Here we show for the first time by genome-wide two-hybrid screens and in vitro binding assays that the PX domain is a bona fide protein interaction domain. The yeast PX domain-only proteins Grd19p (YOR357C) ... and Ypt35p (YHR105W), as well as the isolated PX domains from Mvp1p (YMR004W), Snx42p/Cvt20p/Atg20p (YDL113C), Vam7p (YGL212W), and Vps17p (YOR132W), yielded a total of 40 reproducible two-hybrid interactions. Thirty-five interactions were found for the full-length proteins of Bem1p (YBR200W), Snx42p, Snx4p/Cvt13p (YJL036W), Vam7p, Vps5p (YOR069W), and Vps17p, but these appear not to require the PX domain, because these interactions could not be reproduced with PX-only baits. Interactions of Grd19p, Vam7p, Vps5p, Vps17p, and Ypt35p with members of the Yip1p family of proteins were detected consistently and were verified by in vitro binding assays. The N-terminal cytoplasmic domain of Yip1p and Yif1p mediates these interactions with PX domains. A mutation in the lipid-binding pocket of Ypt35p that reduces lipid binding markedly does not affect these PX domain protein interactions, arguing that lipid binding uses a different interaction surface than protein binding.
Mesh Terms:
Binding Sites, Cloning, Molecular, Mutation, Phosphatidylinositols, Protein Binding, Protein Structure, Tertiary, Proteomics, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Two-Hybrid System Techniques
Binding Sites, Cloning, Molecular, Mutation, Phosphatidylinositols, Protein Binding, Protein Structure, Tertiary, Proteomics, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Two-Hybrid System Techniques
Mol. Cell Proteomics
Date: Nov. 01, 2004
PubMed ID: 15263065
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