Ashcroft, Alison E.; Brinker, Achim; Coyle, Joseph E.; Weber, Frank; Kaiser, Markus; Moroder, Luis; Parsons, Mark R.; Jager, Joachim; Hartl, Ulrich F.; Hayer-Hartl, Manajit; Radford, Sheena E.:
Structural plasticity and non-covalent substrate binding in the GroEL apical domain: A study using electrospray ionisation mass spectrometry and fluorescence binding studies
In: The Journal of Biological Chemistry (JBC), Vol. 277 (2002), No. 36, pp. 33115 - 33126
2002article/chapter in journal
Biology
Related: 1 publication(s)
Title:
Structural plasticity and non-covalent substrate binding in the GroEL apical domain: A study using electrospray ionisation mass spectrometry and fluorescence binding studies
Author:
Ashcroft, Alison E.;Brinker, Achim;Coyle, Joseph E.;Weber, Frank;Kaiser, MarkusUDE
LSF ID
52590
ORCID
0000-0002-6540-8520ORCID iD
Other
connected with university
;
Moroder, Luis;Parsons, Mark R.;Jager, Joachim;Hartl, Ulrich F.;Hayer-Hartl, Manajit;Radford, Sheena E.
Year of publication:
2002

Abstract:

Advances in understanding how GroEL binds to non-native proteins are reported. Conformational flexibility in the GroEL apical domain, which could account for the variety of substrates that GroEL binds, is illustrated by comparison of several independent crystallographic structures of apical domain constructs which show conformational plasticity in helices H and I. Additionally, ESI-MS indicates that apical domain constructs have co-populated conformations at neutral pH. To assess the ability of different apical domain conformers to bind co-chaperone and substrate, model peptides corresponding to the mobile loop of GroES and to helix D from rhodanese were studied. Analysis of apical domain-peptide complexes by ESI-MS indicates that only the folded or partially-folded apical domain conformations form complexes that survive gas phase conditions. Fluorescence binding studies show that the apical domain can fully bind both peptides independently. No competition for binding was observed, suggesting the peptides have distinct apical domain binding sites. Blocking the GroES-apical domain binding site in GroEL rendered the chaperonin inactive in binding GroES and in assisting the folding of denatured rhodanese, but still capable of binding non-native proteins, supporting the conclusion that GroES and substrate proteins have, at least partially, distinct binding sites even in the intact GroEL tetradecamer.