title: Conformational coupling, bridge helix dynamics and active site dehydration in catalysis by RNA polymerase.
abstract: Molecular dynamics simulation of Thermus thermophilus (Tt) RNA polymerase (RNAP) in a catalytic conformation demonstrates that the active site dNMP-NTP base pair must be substantially dehydrated to support full active site closing and optimum conditions for phosphodiester bond synthesis. In silico mutant beta R428A RNAP, which was designed based on substitutions at the homologous position (Rpb2 R512) of Saccharomyces cerevisiae (Sc) RNAP II, was used as a reference structure to compare to Tt RNAP in simulations. Long range conformational coupling linking a dynamic segment of the bridge alpha-helix, the extended fork loop, the active site, and the trigger loop-trigger helix is apparent and adversely affected in beta R428A RNAP. Furthermore, bridge helix bending is detected in the catalytic structure, indicating that bridge helix dynamics may regulate phosphodiester bond synthesis as well as translocation. An active site "latch" assembly that includes a key trigger helix residue Tt beta' H1242 and highly conserved active site residues beta E445 and R557 appears to help regulate active site hydration/dehydration. The potential relevance of these observations in understanding RNAP and DNAP induced fit and fidelity is discussed.
authors.affiliation: Department of Biochemistry and Molecular Biology, Michigan State University, E. Lansing, MI 48824-1319, USA.
authors.forename: Steve A
authors.initials: SA
authors.lastname: Seibold
authors.forename: Badri Nath
authors.initials: BN
authors.lastname: Singh
authors.forename: Chunfen
authors.initials: C
authors.lastname: Zhang
authors.forename: Maria
authors.initials: M
authors.lastname: Kireeva
authors.forename: Céline
authors.initials: C
authors.lastname: Domecq
authors.forename: Annie
authors.initials: A
authors.lastname: Bouchard
authors.forename: Anthony M
authors.initials: AM
authors.lastname: Nazione
authors.forename: Michael
authors.initials: M
authors.lastname: Feig
authors.forename: Robert I
authors.initials: RI
authors.lastname: Cukier
authors.forename: Benoit
authors.initials: B
authors.lastname: Coulombe
authors.forename: Mikhail
authors.initials: M
authors.lastname: Kashlev
authors.forename: Michael
authors.initials: M
authors.lastname: Hampsey
authors.forename: Zachary F
authors.initials: ZF
authors.lastname: Burton
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D008958:Models, Molecular; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D009154:Mutation / Q000235:genetics; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012319:RNA Polymerase II / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D016963:Thermus thermophilus / Q000201:enzymology*
chemical_list: D012319:RNA Polymerase II