TY - JOUR
T1 - Backbone dynamics of Escherichia coli adenylate kinase at the extreme stages of the catalytic cycle studied by 15N NMR relaxation
AU - Shapiro, Yury E.
AU - Sinev, Michael A.
AU - Sineva, Elena V.
AU - Tugarinov, Vitali
AU - Meirovitch, Eva
PY - 2000/6/6
Y1 - 2000/6/6
N2 - 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 AP5A-inhibitor-bound AKeco were studied comparatively with 15N 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 (AKeco*AP5A). Fast internal motions affect 93% of the AKeco sites, with pronounced preference for domains AMPbd and LID, and 47% of the AKeco*AP5A sites, with limited variability along the chain. The mean squared generalized order parameters, (S2), 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.
AB - 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 AP5A-inhibitor-bound AKeco were studied comparatively with 15N 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 (AKeco*AP5A). Fast internal motions affect 93% of the AKeco sites, with pronounced preference for domains AMPbd and LID, and 47% of the AKeco*AP5A sites, with limited variability along the chain. The mean squared generalized order parameters, (S2), 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.
UR - http://www.scopus.com/inward/record.url?scp=0034612241&partnerID=8YFLogxK
U2 - 10.1021/bi992076h
DO - 10.1021/bi992076h
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C2 - 10828981
AN - SCOPUS:0034612241
SN - 0006-2960
VL - 39
SP - 6634
EP - 6644
JO - Biochemistry
JF - Biochemistry
IS - 22
ER -