2J6T

Ternary complex of Sulfolobus solfataricus Dpo4 DNA polymerase, O6- methylguanine modified DNA, and dATP.

Summary for 2J6T

• Entry DOI: 10.2210/pdb2j6t/pdb
• Related: 1JX4 1JXL 1N48 1N56 1RYR 1RYS 1S0M 1S0N 1S0O 1S10 1S97 1S9F 2AGO 2AGP 2AGQ 2ASD 2ASJ 2ASL 2ATL 2AU0 2BQ3 2BQR 2BQU 2BR0 2C22 2C28 2C2D 2C2E 2C2R 2J6S 2J6U
• Descriptor: DNA POLYMERASE IV, 5'-D(*GP*GP*GP*GP*GP*AP*AP*GP*GP*AP *TP*TP*C)-3', 5'-D(*TP*CP*AP*TP*XP*GP*AP*AP*TP*CP *CP*TP*TP*CP*CP*CP*CP*C)-3', ... (6 entities in total)
• Functional Keywords: transferase-dna complex, mutator protein, dna replication, o6-methylguanine, transferase, metal-binding, translesion dna synthesis, dna-directed dna polymerase, dpo4, dna repair, dna damage, dna-binding, nucleotidyltransferase, transferase/dna
• Biological source: SULFOLOBUS SOLFATARICUS
• Cellular location: Cytoplasm (Probable): Q97W02
• Total number of polymer chains: 3
• Total formula weight: 51196.36

Authors

Irimia, A.,Eoff, R.L.,Guengerich, F.P.,Egli, M. (deposition date: 2006-10-04, release date: 2006-11-22, Last modification date: 2023-12-13)

Primary citation

Eoff, R.L.,Irimia, A.,Egli, M.,Guengerich, F.P.
Sulfolobus Solfataricus DNA Polymerase Dpo4 is Partially Inhibited by "Wobble" Pairing between O6- Methylguanine and Cytosine, But Accurate Bypass is Preferred.
J.Biol.Chem., 282:1456-, 2007

PubMed Abstract: We examined the effect of a single O6-methylguanine (O6-MeG) template residue on catalysis by a model Y family polymerase, Dpo4 from Sulfolobus solfataricus. Mass spectral analysis of Dpo4-catalyzed extension products revealed that the enzyme accurately bypasses O6-MeG, with C being the major product (approximately 70%) and T or A being the minor species (approximately 20% or approximately 10%, respectively), consistent with steady-state kinetic parameters. Transient-state kinetic experiments revealed that kpol, the maximum forward rate constant describing polymerization, for dCTP incorporation opposite O6-MeG was approximately 6-fold slower than observed for unmodified G, and no measurable product was observed for dTTP incorporation in the pre-steady state. The lack of any structural information regarding how O6-MeG paired in a polymerase active site led us to perform x-ray crystallographic studies, which show that "wobble" pairing occurs between C and O6-MeG. A structure containing T opposite O6-MeG was solved, but much of the ribose and pyrimidine base density was disordered, in accordance with a much higher Km,dTTP that drives the difference in efficiency between C and T incorporation. The more stabilized C:O6-MeG pairing reinforces the importance of hydrogen bonding with respect to nucleotide selection within a geometrically tolerant polymerase active site.

PubMed: 17105728
DOI: 10.1074/JBC.M609661200

Experimental method

X-RAY DIFFRACTION (2.6 Å)