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Distinct actions of cis and trans ATP within the double ring of the chaperonin GroEL

Nature volume 388, pages 792–798 (1997)Cite this article

Abstract

The chaperonin GroEL is a double-ring structure with a central cavity in each ring that provides an environment for the efficient folding of proteins1,2,3 when capped by the co-chaperone GroES in the presence of adenine nucleotides4,5,6,7,8. Productive folding of the substrate rhodanese has been observed in cis ternary complexes, where GroES and polypeptide are bound to the same ring, formed with either ATP, ADP or non-hydrolysable ATP analogues2,9, suggesting that the specific requirement for ATP is confined to an action in the trans ring that evicts GroES and polypeptide from the cis side9. We show here, however, that for the folding of malate dehydrogenase and Rubisco there is also an absolute requirement for ATP in the cis ring, as ADP and AMP-PNP are unable to promote folding. We investigated the specific roles of binding and hydrolysis of ATP in the cis and trans rings using mutant forms of GroEL that bind ATP but are defective in its hydrolysis. Binding of ATP and GroES in cis initiated productive folding inside a highly stable GroEL–ATP–GroES complex. To discharge GroES and polypeptide, ATP hydrolysis in the cis ring was required to form a GroEL–ADP–GroES complex with decreased stability, priming the cis complex for release by ATP binding (without hydrolysis) in the trans ring. These observations offer an explanation of why GroEL functions as a double-ring complex.

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Figure 1: ATP is required in cis for the GroEL–GroES-mediated folding of two stringent substrates, Rubisco and MDH.
Figure 2: ATP is required in cis for the GroEL–GroES-mediated folding of two stringent substrates, Rubisco and MDH.
Figure 3: ATP is required in cis for the GroEL–GroES-mediated folding of two stringent substrates, Rubisco and MDH.
Figure 4: ATP is required in cis for the GroEL–GroES-mediated folding of two stringent substrates, Rubisco and MDH.
Figure 5: ATP is required in cis for the GroEL–GroES-mediated folding of two stringent substrates, Rubisco and MDH.
Figure 6: Binding of ATP and GroES to the cis ring of GroEL is sufficient to trigger the release and folding of Rubisco.
Figure 7: Binding of ATP and GroES to the cis ring of GroEL is sufficient to trigger the release and folding of Rubisco.
Figure 8: Binding of ATP and GroES to the cis ring of GroEL is sufficient to trigger the release and folding of Rubisco.
Figure 9: ATP hydrolysis in the cis ring ‘primes’ release of GroES and peptide ligands by trans-sided ATP.
Figure 10: ATP hydrolysis in the cis ring ‘primes’ release of GroES and peptide ligands by trans-sided ATP.
Figure 11: ATP hydrolysis in the cis ring ‘primes’ release of GroES and peptide ligands by trans-sided ATP.
Figure 12: Release of GFP refolded in tetradecameric D398A cis ternary complexes by subsequent addition in trans of ATP but not ADP.
Figure 13: Schematic representation of the action of ATP in cis and trans rings during GroEL–GroES-mediated protein folding.

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Acknowledgements

We thank K. Furtak and J. Sutin for technical assistance, and D. Boisvert for discussion. This work was supported by grants from the NIH. S.G.B. is a fellow of the Wellcome Trust.

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Authors and Affiliations

  1. Howard Hughes Medical Institute,

    Hays S. Rye, Zhaohui Xu, Paul B. Sigler & Arthur L. Horwich

  2. Department of Genetics, School of Medicine,

    Hays S. Rye, Steven G. Burston, Wayne A. Fenton & Arthur L. Horwich

  3. Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, 06510, Connecticut, USA

    Joseph M. Beechem

  4. Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, 37232, Tennessee, USA

    Zhaohui Xu & Paul B. Sigler

Author notes

  1. Correspondence and requests for materials should be addressed to A.L.H.

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    Rye, H., Burston, S., Fenton, W. et al. Distinct actions of cis and trans ATP within the double ring of the chaperonin GroEL. Nature 388, 792–798 (1997). https://doi.org/10.1038/42047

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