Conditions for nucleotide-dependent GroES-GroEL interactions. GroEL sub(14)(GroES sub(7)) sub(2) is favored by an asymmetric distribution of nucleotides

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...

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Published in:The Journal of biological chemistry 1997-10, Vol.272 (43), p.26999-27004
Main Authors: Gorovits, B M, Ybarra, J, Seale, J W, Horowitz, P M
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Ybarra, J
Seale, J W
Horowitz, P M
description 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 beta , gamma -imidodiphosphate (AMP-PNP), was investigated by using the fluorescence anisotropy of succinimidyl-1-pyrenebutyrate-labeled GroES. In the presence of Mg super(2+)-ATP and high [KCl] (10 mM), two GroES sub(7) rings bind per one GroEL sub(14). In contrast, in the presence of ADP or AMP-PNP only one molecule of oligomeric GroES can be tightly bound by GroEL. With AMP-PNP, binding of a small amount (
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GroEL sub(14)(GroES sub(7)) sub(2) is favored by an asymmetric distribution of nucleotides</title><source>ScienceDirect Journals</source><creator>Gorovits, B M ; Ybarra, J ; Seale, J W ; Horowitz, P M</creator><creatorcontrib>Gorovits, B M ; Ybarra, J ; Seale, J W ; Horowitz, P M</creatorcontrib><description>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 beta , gamma -imidodiphosphate (AMP-PNP), was investigated by using the fluorescence anisotropy of succinimidyl-1-pyrenebutyrate-labeled GroES. In the presence of Mg super(2+)-ATP and high [KCl] (10 mM), two GroES sub(7) rings bind per one GroEL sub(14). In contrast, in the presence of ADP or AMP-PNP only one molecule of oligomeric GroES can be tightly bound by GroEL. With AMP-PNP, binding of a small amount (&lt;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 GroEL sub(14)(GroES sub(7)) 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 GroEL sub(14) (GroES sub(7)) complex formed in the presence of AMP-PNP. This is the first demonstration of an asymmetric distribution of nucleotides bound on the 1:2 GroEL sub(14) (GroES sub(7)) sub(2) complex. 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title Conditions for nucleotide-dependent GroES-GroEL interactions. GroEL sub(14)(GroES sub(7)) sub(2) is favored by an asymmetric distribution of nucleotides
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