Characterisation of a GroEL Single-Ring Mutant that Supports Growth of Escherichia coli and Has GroES-Dependent ATPase Activity

JOURNAL OF MOLECULAR BIOLOGY

卷:396

Article

Kovacs, Eszter^1,2  Sun, Zhe^1,3  Liu, Han^1,4  Scott, David J.⁵  Karsisiotis, Andreas I.^1,6  Clarke, Anthony R.⁷  Burston, Steven G.⁷  Lund, Peter A.¹ 

[1] Univ Birmingham, Sch Biosci, Birmingham B15 2TT, W Midlands, England

[2] SOLVO Biotechnol, H-2040 Budaors, Hungary

[3] Univ Cambridge, Dept Biochem, Cambridge CB2 1QW, England

[4] Univ Liverpool, Physiol Lab, Liverpool L69 3BX, Merseyside, England

[5] Univ Nottingham, Sch Biosci, Natl Ctr Macromol Hydrodynam, Loughborough LE12 5RD, Leics, England

[6] CNRS, UPR 4301, Ctr Biophys Mol, F-45071 Orleans 2, France

[7] Univ Bristol, Sch Med Sci, Dept Biochem, Bristol BS8 1TD, Avon, England

关键词: chaperonin;    GroEL;    single ring;    allostery;   

DOI  :  10.1016/j.jmb.2009.11.074

来源: Elsevier

【 摘 要 】

Binding and folding of substrate proteins by the molecular chaperone GroEL alternates between its two seven-membered rings in an ATP-regulated manner. The association of ATP and GroES to a polypeptide-bound ring of GroEL encapsulates the folding proteins in the central cavity of that ring (cis ring) and allows it to fold in a protected environment where the risk of aggregation is reduced. ATP hydrolysis in the cis ring changes the potentials within the system such that ATP binding to the opposite (trans) ring triggers the release of all ligands from the cis ring of GroEL through a complex network of allosteric communication between the rings. Inter-ring allosteric communication thus appears indispensable for the function of GroEL, and an engineered single-ring version (SR1) cannot substitute for GroEL in vivo. We describe here the isolation and characterisation of an active single-ring form of the GroEL protein (SR-A92T), which has an exceptionally low ATPase activity that is strongly stimulated by the addition of GroES. Dissection of the kinetic pathway of the ATP-induced structural changes in this active single ring can be explained by the fact that the mutation effectively blocks progression through the full allosteric pathway of the GroEL reaction cycle, thus trapping an early allosteric intermediate. Addition of GroES is able to overcome this block by binding this intermediate and pulling the allosteric pathway to completion via mass action, explaining how bacterial cells expressing this protein as their only chaperonin are viable. (C) 2009 Elsevier Ltd. All rights reserved.

【 授权许可】

Free