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Title: Combined Use of Residual Dipolar Couplings and Solution X-ray Scattering To Rapidly Probe Rigid-Body Conformational Transitions in a Non-phosphorylatable Active-Site Mutant of the 128 kDa Enzyme I Dimer

Abstract

The first component of the bacterial phosphotransferase system, enzyme I (EI), is a multidomain 128 kDa dimer that undergoes large rigid-body conformational transitions during the course of its catalytic cycle. Here we investigate the solution structure of a non-phosphorylatable active-site mutant in which the active-site histidine is substituted by glutamine. We show that perturbations in the relative orientations and positions of the domains and subdomains can be rapidly and reliably determined by conjoined rigid-body/torsion angle/Cartesian simulated annealing calculations driven by orientational restraints from residual dipolar couplings and shape and translation information afforded by small- and wide-angle X-ray scattering. Although histidine and glutamine are isosteric, the conformational space available to a Gln side chain is larger than that for the imidazole ring of His. An additional hydrogen bond between the side chain of Gln189 located on the EIN{sup {alpha}/{beta}} subdomain and an aspartate (Asp129) on the EIN{sup {alpha}} subdomain results in a small ({approx}9{sup o}) reorientation of the EIN{sup {alpha}} and EIN{sup {alpha}/{beta}} subdomains that is in turn propagated to a larger reorientation ({approx}26{sup o}) of the EIN domain relative to the EIC dimerization domain, illustrating the positional sensitivity of the EIN domain and its constituent subdomains to small structural perturbations.

Authors:
; ; ; ;  [1]
  1. (NIH)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
OTHERNIH
OSTI Identifier:
1050751
Resource Type:
Journal Article
Journal Name:
J. Am. Chem. Soc.
Additional Journal Information:
Journal Volume: 133; Journal Issue: (3) ; 01, 2011; Journal ID: ISSN 0002-7863
Country of Publication:
United States
Language:
ENGLISH
Subject:
08 HYDROGEN; ANNEALING; COUPLINGS; DIMERIZATION; DIMERS; ENZYMES; GLUTAMINE; HISTIDINE; HYDROGEN; IMIDAZOLES; MUTANTS; PHOSPHOTRANSFERASES; RESTRAINTS; SCATTERING; SENSITIVITY; SHAPE; SOLUTIONS

Citation Formats

Takayama, Yuki, Schwieters, Charles D., Grishaev, Alexander, Ghirlando, Rodolfo, and Clore, G. Marius. Combined Use of Residual Dipolar Couplings and Solution X-ray Scattering To Rapidly Probe Rigid-Body Conformational Transitions in a Non-phosphorylatable Active-Site Mutant of the 128 kDa Enzyme I Dimer. United States: N. p., 2012. Web. doi:10.1021/ja109866w.
Takayama, Yuki, Schwieters, Charles D., Grishaev, Alexander, Ghirlando, Rodolfo, & Clore, G. Marius. Combined Use of Residual Dipolar Couplings and Solution X-ray Scattering To Rapidly Probe Rigid-Body Conformational Transitions in a Non-phosphorylatable Active-Site Mutant of the 128 kDa Enzyme I Dimer. United States. doi:10.1021/ja109866w.
Takayama, Yuki, Schwieters, Charles D., Grishaev, Alexander, Ghirlando, Rodolfo, and Clore, G. Marius. Tue . "Combined Use of Residual Dipolar Couplings and Solution X-ray Scattering To Rapidly Probe Rigid-Body Conformational Transitions in a Non-phosphorylatable Active-Site Mutant of the 128 kDa Enzyme I Dimer". United States. doi:10.1021/ja109866w.
@article{osti_1050751,
title = {Combined Use of Residual Dipolar Couplings and Solution X-ray Scattering To Rapidly Probe Rigid-Body Conformational Transitions in a Non-phosphorylatable Active-Site Mutant of the 128 kDa Enzyme I Dimer},
author = {Takayama, Yuki and Schwieters, Charles D. and Grishaev, Alexander and Ghirlando, Rodolfo and Clore, G. Marius},
abstractNote = {The first component of the bacterial phosphotransferase system, enzyme I (EI), is a multidomain 128 kDa dimer that undergoes large rigid-body conformational transitions during the course of its catalytic cycle. Here we investigate the solution structure of a non-phosphorylatable active-site mutant in which the active-site histidine is substituted by glutamine. We show that perturbations in the relative orientations and positions of the domains and subdomains can be rapidly and reliably determined by conjoined rigid-body/torsion angle/Cartesian simulated annealing calculations driven by orientational restraints from residual dipolar couplings and shape and translation information afforded by small- and wide-angle X-ray scattering. Although histidine and glutamine are isosteric, the conformational space available to a Gln side chain is larger than that for the imidazole ring of His. An additional hydrogen bond between the side chain of Gln189 located on the EIN{sup {alpha}/{beta}} subdomain and an aspartate (Asp129) on the EIN{sup {alpha}} subdomain results in a small ({approx}9{sup o}) reorientation of the EIN{sup {alpha}} and EIN{sup {alpha}/{beta}} subdomains that is in turn propagated to a larger reorientation ({approx}26{sup o}) of the EIN domain relative to the EIC dimerization domain, illustrating the positional sensitivity of the EIN domain and its constituent subdomains to small structural perturbations.},
doi = {10.1021/ja109866w},
journal = {J. Am. Chem. Soc.},
issn = {0002-7863},
number = (3) ; 01, 2011,
volume = 133,
place = {United States},
year = {2012},
month = {10}
}