Crystal structure of the Sec4p.Sec2p complex in the nucleotide exchanging intermediate state

Proc Natl Acad Sci U S A. 2007 May 15;104(20):8305-10. doi: 10.1073/pnas.0701550104. Epub 2007 May 8.

Abstract

Vesicular transport during exocytosis is regulated by Rab GTPase (Sec4p in yeast), which is activated by a guanine nucleotide exchange factor (GEF) called Sec2p. Here, we report the crystal structure of the Sec2p GEF domain in a complex with the nucleotide-free Sec4p at 2.7 A resolution. Upon complex formation, the Sec2p helices approach each other, flipping the side chain of Phe-109 toward Leu-104 and Leu-108 of Sec2p. These three residues provide a hydrophobic platform to attract the side chains of Phe-49, Ile-53, and Ile-55 in the switch I region as well as Phe-57 and Trp-74 in the interswitch region of Sec4p. Consequently, the switch I and II regions are largely deformed, to create a flat hydrophobic interface that snugly fits the surface of the Sec2p coiled coil. These drastic conformational changes disrupt the interactions between switch I and the bound guanine nucleotide, which facilitates the GDP release. Unlike the recently reported 3.3 A structure of the Sec4p.Sec2p complex, our structure contains a phosphate ion bound to the P-loop, which may represent an intermediate state of the nucleotide exchange reaction.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Crystallography, X-Ray
  • Dimerization
  • GTP-Binding Proteins / chemistry*
  • Guanine Nucleotide Exchange Factors / chemistry*
  • Molecular Sequence Data
  • Protein Binding
  • Protein Structure, Secondary
  • Saccharomyces cerevisiae / chemistry*
  • Saccharomyces cerevisiae Proteins / chemistry*
  • rab GTP-Binding Proteins / chemistry*

Substances

  • Guanine Nucleotide Exchange Factors
  • SEC2 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • GTP-Binding Proteins
  • SEC4 protein, S cerevisiae
  • rab GTP-Binding Proteins

Associated data

  • PDB/2EQB