title: Backbone dynamics of escherichia coli adenylate kinase at the extreme stages of the catalytic cycle studied by (15)N NMR relaxation.
abstract: Adenylate kinase from Escherichia coli (AKeco), consisting of a single 23.6 kDa polypeptide chain folded into domains CORE, AMPbd, and LID, catalyzes the reaction AMP + ATP --> 2ADP. Domains LID and AMPbd execute large-scale movements during catalysis. Backbone dynamics of ligand-free and AP(5)A-inhibitor-bound AKeco were studied comparatively with (15)N NMR relaxation methods. Overall diffusion with correlation times of 15.05 (11.42) ns and anisotropy D(parallel)/D(perp) = 1.25 (1.10), and fast internal motions with correlation times up to 100 ps (50 ps), were determined for AKeco (AKecoAP(5)A). Fast internal motions affect 93% of the AKeco sites, with pronounced preference for domains AMPbd and LID, and 47% of the AKecoAP(5)A sites, with limited variability along the chain. The mean squared generalized order parameters, <S(2)>, of secondary structure elements and loops are affected by ligand binding differentially and in a domain-specific manner. Nanosecond motions predominate within AMPbd. Prominent exchange contributions, associated in particular with residue G10 of the nucleotide-binding P-loop motif, are interpreted to reflect hydrogen-bond dynamics at the inhibitor-binding site. The hypothesis of energetic counter balancing of substrate binding based on crystallographic data is strongly supported by the solution NMR results. Correlations between backbone dynamics and domain displacement are established.
authors.affiliation: Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan 52900, Israel, Department of Structural Biology, The Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Y E
authors.initials: YE
authors.lastname: Shapiro
authors.forename: M A
authors.initials: MA
authors.lastname: Sinev
authors.forename: E V
authors.initials: EV
authors.lastname: Sineva
authors.forename: V
authors.initials: V
authors.lastname: Tugarinov
authors.forename: E
authors.initials: E
authors.lastname: Meirovitch
mesh_terms: D000263:Adenylate Kinase / Q000737:chemistry*; D004058:Diffusion; D015226:Dinucleoside Phosphates / Q000737:chemistry; D004791:Enzyme Inhibitors / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology*; D006860:Hydrogen Bonding; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D017433:Protein Structure, Secondary
chemical_list: D015226:Dinucleoside Phosphates; D004791:Enzyme Inhibitors; C012275:P(1),P(5)-di(adenosine-5'-)pentaphosphate; D000263:Adenylate Kinase