title: Rate-limiting domain and loop motions in arginine kinase.
abstract: Arginine kinase catalyzes the reversible transfer of a phosphoryl group between ATP and arginine. It is the arthropod homologue of creatine kinase, buffering cellular ATP levels. Crystal structures of arginine kinase, in substrate-free and substrate-bound forms, have revealed large conformational changes associated with the catalytic cycle. Recent nuclear magnetic resonance identified movements of the N-terminal domain and a loop comprising residues I182--G209 with conformational exchange rates in the substrate-free enzyme similar to the turnover rate. Here, to understand whether these motions might be rate-limiting, we determined activation barriers for both the intrinsic dynamics and enzyme turnover using measurements over a temperature range of 15-30 °C. (15)N transverse relaxation dispersion yields activation barriers of 46 ± 8 and 34 ± 12 kJ/mol for the N-terminal domain and I182--G209 loop, respectively. An activation barrier of 34 ± 13 kJ/mol was obtained for enzyme turnover from steady-state kinetics. The similarity between the activation barriers is indeed consistent with turnover being limited by backbone conformational dynamics and pinpoints the locations of potentially rate-limiting motions.
authors.affiliation: Department of Biochemistry and Molecular Biology, Oregon Health and Science University, Portland, Oregon 97239-3098, United States.
authors.forename: Omar
authors.initials: O
authors.lastname: Davulcu
authors.forename: Jack J
authors.initials: JJ
authors.lastname: Skalicky
authors.forename: Michael S
authors.initials: MS
authors.lastname: Chapman
mesh_terms: D000818:Animals; D001122:Arginine Kinase / Q000737:chemistry* / Q000378:metabolism*; D020134:Catalytic Domain; D006737:Horseshoe Crabs / Q000201:enzymology*; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013997:Time Factors
chemical_list: D001122:Arginine Kinase