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PDBsum entry 2m2t
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References listed in PDB file
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Key reference
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Title
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How a low-Fidelity DNA polymerase chooses non-Watson-Crick from watson-Crick incorporation.
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Authors
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W.J.Wu,
M.I.Su,
J.L.Wu,
S.Kumar,
L.H.Lim,
C.W.Wang,
F.H.Nelissen,
M.C.Chen,
J.F.Doreleijers,
S.S.Wijmenga,
M.D.Tsai.
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Ref.
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J Am Chem Soc, 2014,
136,
4927-4937.
[DOI no: ]
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PubMed id
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Abstract
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A dogma for DNA polymerase catalysis is that the enzyme binds DNA first,
followed by MgdNTP. This mechanism contributes to the selection of correct dNTP
by Watson-Crick base pairing, but it cannot explain how low-fidelity DNA
polymerases overcome Watson-Crick base pairing to catalyze non-Watson-Crick dNTP
incorporation. DNA polymerase X from the deadly African swine fever virus (Pol
X) is a half-sized repair polymerase that catalyzes efficient dG:dGTP
incorporation in addition to correct repair. Here we report the use of solution
structures of Pol X in the free, binary (Pol X:MgdGTP), and ternary (Pol
X:DNA:MgdGTP with dG:dGTP non-Watson-Crick pairing) forms, along with functional
analyses, to show that Pol X uses multiple unprecedented strategies to achieve
the mutagenic dG:dGTP incorporation. Unlike high fidelity polymerases, Pol X can
prebind purine MgdNTP tightly and undergo a specific conformational change in
the absence of DNA. The prebound MgdGTP assumes an unusual syn conformation
stabilized by partial ring stacking with His115. Upon binding of a gapped DNA,
also with a unique mechanism involving primarily helix αE, the prebound
syn-dGTP forms a Hoogsteen base pair with the template anti-dG. Interestingly,
while Pol X prebinds MgdCTP weakly, the correct dG:dCTP ternary complex is
readily formed in the presence of DNA. H115A mutation disrupted MgdGTP binding
and dG:dGTP ternary complex formation but not dG:dCTP ternary complex formation.
The results demonstrate the first solution structural view of DNA polymerase
catalysis, a unique DNA binding mode, and a novel mechanism for non-Watson-Crick
incorporation by a low-fidelity DNA polymerase.
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