Possible roles of Mg2 ion in Hammerhead RNA inferred from molecular dynamics simulations

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Possible roles of Mg2 ion in Hammerhead RNA
inferred from molecular dynamics simulations

• Taisung Lee
• 7/20/06 York Group Meeting

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Outline

• Hammer Head RNA
• The problems
• The opportunities
• Recent results

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Hammer Head RNA

• The serine protease of RNA enzymes
• The known smallest (and hence the easiest?) RNA
  enzyme so far.
• Cut RNA into segments in the rolling circle
  replication.

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Minimal Hammerhead RNA

• The minimal sequence (43 res.) that is
  catalytically functional.
• In crystal gt in water (5X)
• Lots of studies in vitro and many structures
  available, but not many successful studies in
  silico.

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The problems for current available structures and
data

• Totally conserved A9 and G8.
• Large conformational change(s) necessary?
• When before the reaction? Or occurring at the
  same time?
• What are the general base and acid (pH
  dependence, peaked at 8.5)?
• Roles of Mg2 ions at various positions
• Thio-substitution results

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The opportunities

• Full length (63 res.) X-Ray structure from Bill
  Scott at UCSC.
• More effective than minimal Hammerhead RNA in
  crystal
• Conformation
• almost perfect inline position for the reaction.
• G8 and A9 are nearby now.

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Recent simulations

• RHDO water box. 30,000 atoms totally.
• Total 1 ns water and ion equilibration.
• A Mg2 was artificially added in the middle of
  O2Ps of A9 and scissile site.

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Recent simulations

• Reactant
• Early-TS mimic (Evelyns parameter)
• Late-TS mimic (modified OPO distances, modified
  charge on the leaving group O)
• 3ns production run for each.
• 27 ps QM/MM simulation

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Mg2 roles

• Holds two OP2 atoms hence the overall structure
• MgO2(G8) Possibly causes the pKa shift of O2
  of G8
• MgO5(C1.1) Possibly stabilizes the leaving
  group.
• Helps proton transfer from O2(G8) to O5 (C1.1)

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Next

• Longer simulations
• Simulations without Mg2
• QM/MM simulation along the reaction coordinate.
• Mutation effect
• Thio-substitution effect