title: The energy landscape of adenylate kinase during catalysis.
abstract: Kinases perform phosphoryl-transfer reactions in milliseconds; without enzymes, these reactions would take about 8,000 years under physiological conditions. Despite extensive studies, a comprehensive understanding of kinase energy landscapes, including both chemical and conformational steps, is lacking. Here we scrutinize the microscopic steps in the catalytic cycle of adenylate kinase, through a combination of NMR measurements during catalysis, pre-steady-state kinetics, molecular-dynamics simulations and crystallography of active complexes. We find that the Mg(2+) cofactor activates two distinct molecular events: phosphoryl transfer (>10(5)-fold) and lid opening (10(3)-fold). In contrast, mutation of an essential active site arginine decelerates phosphoryl transfer 10(3)-fold without substantially affecting lid opening. Our results highlight the importance of the entire energy landscape in catalysis and suggest that adenylate kinases have evolved to activate key processes simultaneously by precise placement of a single, charged and very abundant cofactor in a preorganized active site.
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: S Jordan
authors.initials: SJ
authors.lastname: Kerns
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Roman V
authors.initials: RV
authors.lastname: Agafonov
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Young-Jin
authors.initials: YJ
authors.lastname: Cho
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Francesco
authors.initials: F
authors.lastname: Pontiggia
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Renee
authors.initials: R
authors.lastname: Otten
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Dimitar V
authors.initials: DV
authors.lastname: Pachov
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Steffen
authors.initials: S
authors.lastname: Kutter
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Lien A
authors.initials: LA
authors.lastname: Phung
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Padraig N
authors.initials: PN
authors.lastname: Murphy
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Vu
authors.initials: V
authors.lastname: Thai
authors.forename: Tom
authors.initials: T
authors.lastname: Alber
authors.affiliation: Department of Physics, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Michael F
authors.initials: MF
authors.lastname: Hagan
authors.affiliation: Howard Hughes Medical Institute, Department of Biochemistry, Brandeis University, Waltham, Massachusetts, USA.
authors.forename: Dorothee
authors.initials: D
authors.lastname: Kern
mesh_terms: D000263:Adenylate Kinase / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular
chemical_list: D000263:Adenylate Kinase