title: Characterization of the Domain Orientations of E. coli 5'-Nucleotidase by Fitting an Ensemble of Conformers to DEER Distance Distributions.
abstract: Escherichia coli 5'-nucleotidase is a two-domain enzyme exhibiting a unique 96° domain motion that is required for catalysis. Here we present an integrated structural biology study that combines DEER distance distributions with structural information from X-ray crystallography and computational biology to describe the population of presumably almost isoenergetic open and closed states in solution. Ensembles of models that best represent the experimental distance distributions are determined by a Monte Carlo search algorithm. As a result, predominantly open conformations are observed in the unliganded state indicating that the majority of enzyme molecules await substrate binding for the catalytic cycle. The addition of a substrate analog yields ensembles with an almost equal mixture of open and closed states. Thus, in the presence of substrate, efficient catalysis is provided by the simultaneous appearance of open conformers (binding substrate or releasing product) and closed conformers (enabling the turnover of the substrate).
authors.affiliation: Institute of Bioanalytical Chemistry, Center for Biotechnology and Biomedicine, University of Leipzig, Deutscher Platz 5, 04103 Leipzig, Germany.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Krug
authors.affiliation: Center for Structural Biology, Vanderbilt University, Nashville, TN 37212, USA; Department of Chemistry, Vanderbilt University, Nashville, TN 37212, USA.
authors.forename: Nathan S
authors.initials: NS
authors.lastname: Alexander
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37212, USA.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Stein
authors.affiliation: Institute of Bioanalytical Chemistry, Center for Biotechnology and Biomedicine, University of Leipzig, Deutscher Platz 5, 04103 Leipzig, Germany.
authors.forename: Antje
authors.initials: A
authors.lastname: Keim
authors.affiliation: Center for Structural Biology, Vanderbilt University, Nashville, TN 37212, USA; Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37212, USA.
authors.forename: Hassane S
authors.initials: HS
authors.lastname: Mchaourab
authors.affiliation: Institute of Bioanalytical Chemistry, Center for Biotechnology and Biomedicine, University of Leipzig, Deutscher Platz 5, 04103 Leipzig, Germany. Electronic address: strater@bbz.uni-leipzig.de.
authors.forename: Norbert
authors.initials: N
authors.lastname: Sträter
authors.affiliation: Center for Structural Biology, Vanderbilt University, Nashville, TN 37212, USA; Department of Chemistry, Vanderbilt University, Nashville, TN 37212, USA; Vanderbilt University, 7330 Stevenson Center, Station B 351822, Nashville, TN 37235, USA. Electronic address: jens.meiler@vanderbilt.edu.
authors.forename: Jens
authors.initials: J
authors.lastname: Meiler
mesh_terms: D015720:5'-Nucleotidase / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary
chemical_list: D029968:Escherichia coli Proteins; D015720:5'-Nucleotidase