The structure of the Guanine Nucleotide Exchange Factor Rlf in complex with the small G-protein Ral identifies conformational intermediates of the exchange reaction and the basis for the selectivity

J Struct Biol. 2016 Feb;193(2):106-14. doi: 10.1016/j.jsb.2015.12.006. Epub 2015 Dec 11.

Abstract

CDC25 homology domain (CDC25-HD) containing Guanine Nucleotide Exchange Factors (GEFs) initiate signalling by small G-proteins of the Ras-family. Each GEF acts on a small subset of the G-proteins only, thus providing signalling selectivity. Rlf is a GEF with selectivity for the G-proteins RalA and RalB. Here the crystal structure of Rlf in complex with Ral is determined. The Rlf·Ral complex crystallised into two different crystal forms, which represent different steps of the exchange reaction. Thereby general insight in the CDC25-HD catalysed nucleotide exchange is obtained. In addition, the basis for the selectivity of the interaction is investigated. The exchange activity is monitored by the use of recombinant proteins. Selectivity determinants in the binding interface are identified and confirmed by a mutational study.

Keywords: CDC25-homology domain; Guanine nucleotide exchange reaction; Induced fit; Ras-family of G-proteins.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallography, X-Ray
  • Drosophila Proteins / chemistry*
  • Drosophila Proteins / genetics
  • Drosophila Proteins / metabolism
  • Guanine Nucleotide Exchange Factors
  • Multiprotein Complexes / chemistry
  • Multiprotein Complexes / metabolism
  • Protein Conformation
  • Transcription Factors / chemistry*
  • Transcription Factors / genetics
  • Transcription Factors / metabolism
  • ral GTP-Binding Proteins / chemistry*
  • ral GTP-Binding Proteins / genetics
  • ral GTP-Binding Proteins / metabolism

Substances

  • Drosophila Proteins
  • Guanine Nucleotide Exchange Factors
  • Multiprotein Complexes
  • Ralb protein, human
  • Rgl2 protein, mouse
  • Transcription Factors
  • ral GTP-Binding Proteins