title: Mg2+-assisted catalysis by B. stearothermophilus TrpRS is promoted by allosteric effects.
abstract: Mn(2+)-assisted catalysis by B. stearothermophilus TrpRS parallels that in polymerases and reduces specificity in amino acid activation. As predicted by nonequilibrium molecular dynamics simulations, multivariant thermodynamic cycles with [ATP]-dependent Michaelis-Menten kinetics and Mn(2+) for Mg(2+) substitution demonstrate energetic coupling of ATP affinities to the metal; to lysines K111 and K192, which interact via the PPi leaving group; and to K195, which couples differently to the metal via the alpha-phosphate. However, net coupling to the metal opposes catalysis in both ground (K(M)) and transition (k(cat)) states. The 10(5)-fold rate acceleration by Mg(2+)-protein interactions therefore requires additional favorable protein-metal couplings. Examples include longer-range, i.e., allosteric, interactions previously illustrated by the remote F37I mutation, which both reduces k(cat) and enhances catalytic assist by Mn(2+), relative to that by Mg(2+). These data support a model linking metal-assisted phosphoryl transfer catalysis to domain movement, and hence to free-energy transduction in a broad range of enzymes.
authors: Department of Biochemistry and Biophysics, CB 7260, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7260, USA.
authors: Violetta
authors: V
authors: Weinreb
authors: Li
authors: L
authors: Li
authors: Cassandra L
authors: CL
authors: Campbell
authors: Laurie S
authors: LS
authors: Kaguni
authors: Charles W
authors: CW
authors: Carter
mesh_terms: D000494:Allosteric Regulation; D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000378:metabolism; D007700:Kinetics; D008239:Lysine / Q000737:chemistry; D008274:Magnesium / Q000737:chemistry* / Q000378:metabolism; D008670:Metals / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013816:Thermodynamics; D014369:Tryptophan-tRNA Ligase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D008670:Metals; D014369:Tryptophan-tRNA Ligase; D008274:Magnesium; D008239:Lysine