Conditions for nucleotide-dependent GroES-GroEL interactions. GroEL14(GroES7)2 is favored by an asymmetric distribution of nucleotides

Boris M. Gorovits, Jesse Ybarra, Jeffrey W. Seale, Paul M. Horowitz

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

A still unresolved question regarding the mechanism of chaperonin- assisted protein folding involves the stoichiometry of the GroEL-GroES complex. This is important, because the activities of the Escherichia coli chaperonin GroEL are modulated by the cochaperonin GroES. In this report, the binding of GroES to highly purified GroEL in the presence of ATP, ADP, and the nonhydrolyzable ATP analogue, 5'-adenylyl β,γ-imido-diphosphate (AMP- PNP), was investigated by using the fluorescence anisotropy of succinimidyl- 1-pyrenebutyrate-labeled GroES. In the presence of Mg2+-ATP and high [KCI] (10 mM), two GroES7 rings bind per one GroEL14. In contrast, in the presence of ADP or AMPPNP only one molecule of oligomeric GroES can be tightly bound by GroEL. With AMP-PNP, binding of a small amount (<20%) of a second GroES can be detected. In the presence of ADP alone, a second GroES ring can bind to GroEL weakly and with negative cooperativity. Strikingly, addition of AMP-PNP to the solution containing preformed GroEL14/(GroES7) complexes formed in the presence of ADP results in an increase in the fluorescence anisotropy. Analysis of this effect indicates that 2 mol of GroES oligomer can be bound in the presence of mixed nucleotides. A similar conclusion follows from studies in which ADP is added to an GroEL14 (GroES7) complex formed in the presence of AMP-PNP. This is the first demonstration of an asymmetric distribution of nucleotides bound on the 1:2 GroEL14 (GroES7)2 complex. The relation of the observed phenomena to the proposed mechanism of the GroEL function is discussed.

Original languageEnglish (US)
Pages (from-to)26999-27004
Number of pages6
JournalJournal of Biological Chemistry
Volume272
Issue number43
DOIs
StatePublished - 1997
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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