2w9c
Ternary complex of Dpo4 bound to N2,N2-dimethyl-deoxyguanosine modified DNA with incoming dTTP
Structural highlights
FunctionDPO4_SACS2 Poorly processive, error-prone DNA polymerase involved in untargeted mutagenesis. Copies undamaged DNA at stalled replication forks, which arise in vivo from mismatched or misaligned primer ends. These misaligned primers can be extended by PolIV. Exhibits no 3'-5' exonuclease (proofreading) activity. It is involved in translesional synthesis. Evolutionary Conservation Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf. Publication Abstract from PubMedPrevious work has shown that Y-family DNA polymerases tolerate large DNA adducts, but a substantial decrease in catalytic efficiency and fidelity occurs during bypass of N(2),N(2)-dimethyl (Me(2))-substituted guanine (N(2),N(2)-Me(2)G), in contrast to a single methyl substitution. Therefore, it is unclear why the addition of two methyl groups is so disruptive. The presence of N(2),N(2)-Me(2)G lowered the catalytic efficiency of the model enzyme Sulfolobus solfataricus Dpo4 16,000-fold. Dpo4 inserted dNTPs almost at random during bypass of N(2),N(2)-Me(2)G, and much of the enzyme was kinetically trapped by an inactive ternary complex when N(2),N(2)-Me(2)G was present, as judged by a reduced burst amplitude (5% of total enzyme) and kinetic modeling. One crystal structure of Dpo4 with a primer having a 3'-terminal dideoxycytosine (C(dd)) opposite template N(2),N(2)-Me(2)G in a post-insertion position showed C(dd) folded back into the minor groove, as a catalytically incompetent complex. A second crystal had two unique orientations for the primer terminal C(dd) as follows: (i) flipped into the minor groove and (ii) a long pairing with N(2),N(2)-Me(2)G in which one hydrogen bond exists between the O-2 atom of C(dd) and the N-1 atom of N(2),N(2)-Me(2)G, with a second water-mediated hydrogen bond between the N-3 atom of C(dd) and the O-6 atom of N(2),N(2)-Me(2)G. A crystal structure of Dpo4 with dTTP opposite template N(2),N(2)-Me(2)G revealed a wobble orientation. Collectively, these results explain, in a detailed manner, the basis for the reduced efficiency and fidelity of Dpo4-catalyzed bypass of N(2),N(2)-Me(2)G compared with mono-substituted N(2)-alkyl G adducts. Structure-Function Relationships in Miscoding by Sulfolobus solfataricus DNA Polymerase Dpo4: GUANINE N2,N2-DIMETHYL SUBSTITUTION PRODUCES INACTIVE AND MISCODING POLYMERASE COMPLEXES.,Zhang H, Eoff RL, Kozekov ID, Rizzo CJ, Egli M, Guengerich FP J Biol Chem. 2009 Jun 26;284(26):17687-99. PMID:19542237[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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