title: Evidence for a hydroxide ion bridging two magnesium ions at the active site of the hammerhead ribozyme.
abstract: In the presence of magnesium ions, cleavage by the hammerhead ribozyme RNA at a specific residue leads to 2'3'-cyclic phosphate and 5'-OH extremities. In the cleavage reaction an activated ribose 2'-hydroxyl group attacks its attached 3'-phosphate. Molecular dynamics simulations of the crystal structure of the hammerhead ribozyme, obtained after flash-freezing of crystals under conditions where the ribozyme is active, provide evidence that a mu-bridging OH-ion is located between two Mg2+ions close to the cleavable phosphate. Constrained simulations show further that a flip from the C3'- endo to the C2'- endo conformation of the ribose at the cleavable phosphate brings the 2'-hydroxyl in proximity to both the attacked phosphorous atom and the mu-bridging OH-ion. Thus, the simulations lead to a detailed new insight into the mechanism of hammerhead ribozyme cleavage where a mu-hydroxo bridged magnesium cluster, located on the deep groove side, provides an OH-ion that is able to activate the 2'-hydroxyl nucleophile after a minor and localized conformational change in the RNA.
authors.affiliation: Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 rue René Descartes, F-67084 Strasbourg, France.
authors.forename: T
authors.initials: T
authors.lastname: Hermann
authors.forename: P
authors.initials: P
authors.lastname: Auffinger
authors.forename: W G
authors.initials: WG
authors.lastname: Scott
authors.forename: E
authors.initials: E
authors.lastname: Westhof
mesh_terms: D001665:Binding Sites; D003460:Crystallization; D004563:Electrochemistry; D005615:Freezing; D006878:Hydroxides / Q000737:chemistry*; D008274:Magnesium / Q000737:chemistry*; D008958:Models, Molecular; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D016337:RNA, Catalytic / Q000737:chemistry* / Q000378:metabolism
chemical_list: D006878:Hydroxides; D016337:RNA, Catalytic; C031356:hydroxide ion; D008274:Magnesium