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Ionic interactions at both inter‐ring contact sites of GroEL are involved in transmission of the allosteric signal: A time‐resolved infrared difference study
Author(s) -
Sot Begoña,
von Germar Fritzthof,
Mäntele Werner,
Valpuesta Jose María,
Taneva Stefka G.,
Muga Arturo
Publication year - 2005
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1110/ps.051469605
Subject(s) - allosteric regulation , groel , ring (chemistry) , chemistry , cooperativity , biophysics , crystallography , ionic bonding , protein folding , chaperonin , biochemistry , biology , ion , escherichia coli , organic chemistry , gene , enzyme
The biological activity of the double‐ring chaperonin GroEL is regulated by complex allosteric interactions, which include positive intra‐ring and negative inter‐ring cooperativity. To further characterize inter‐ring communication, the nucleotide‐induced absorbance changes in the vibrational spectrum of the chaperonin GroEL, of two single‐point mutants suppressing one inter‐ring ionic contact (E461K and E434K) and of a single‐ring version of this protein, were investigated by time‐resolved infrared difference spectroscopy. Interaction of the nucleotide with the proteins was triggered by its photochemical release from a biologically inactive caged precursor [P 3 ‐1‐(2‐nitro) phenylethyl nucleotide]. The results indicate that (1) ATP binding to the protein induces a conformational change that affects concomitantly both intra‐ring and inter‐ring communication, and (2) the experimental absorbance changes are sensitive to the double‐ring structure of the protein. The characterization of the single‐point, inter‐ring mutants demonstrates that ionic interactions at both contact sites are involved in the transmission of the allosteric signal. However, both mutations have different effects on the inter‐ring interface. While that of E461K still retains ionic contacts sensitive to ATP binding, E434K shows spectroscopic features similar to those of the single‐ring version of the protein, therefore suggesting that electrostatic interactions at these contact sites contribute differently to the stability of the inter‐ring interface.