2JEI

The Molecular Basis of Selectivity of Nucleoside Triphosphate Incorporation Opposite O6-Benzylguanine by Sulfolobus solfataricus DNA Polymerase IV: Steady-state and Pre-steady-state Kinetics and X- Ray Crystallography of Correct and Incorrect Pairing

Summary for 2JEI

• Entry DOI: 10.2210/pdb2jei/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 2J6T 2J6U 2JEF 2JEG 2JEJ
• Descriptor: DNA POLYMERASE IV, 5'-D(*GP*GP*GP*GP*GP*AP*AP*GP*GP*AP *TP*TP*CP*T)-3', 5'-D(*TP*CP*AP*C BZGP*GP*AP*AP*TP*CP*CP *TP*TP*CP*CP*CP*CP*C)-3', ... (6 entities in total)
• Functional Keywords: dna replication, mutator protein, 06-benzylguanine, nucleotidyltransferase, dna-directed dna polymerase, dpo4, magnesium, polymerase, dna damage, dna repair, alkylating agents, translesion synthesis, transferase, dna-binding, metal-binding
• Biological source: SULFOLOBUS SOLFATARICUS
• Total number of polymer chains: 3
• Total formula weight: 51617.71

Authors

Eoff, R.L.,Angel, K.C.,Kosekov, I.D.,Egli, M.,Guengerich, F.P. (deposition date: 2007-01-17, release date: 2007-03-13, Last modification date: 2023-12-13)

Primary citation

Eoff, R.L.,Angel, K.C.,Egli, M.,Guengerich, F.P.
Molecular Basis of Selectivity of Nucleoside Triphosphate Incorporation Opposite O6-Benzylguanine by Sulfolobus Solfataricus DNA Polymerase Dpo4: Steady-State and Pre-Steady-State Kinetics and X-Ray Crystallography of Correct and Incorrect Pairing.
J.Biol.Chem., 282:13573-, 2007

PubMed Abstract: Previous work has shown that Sulfolobus solfataricus DNA polymerase Dpo4-catalyzed bypass of O(6)-methylguanine (O(6)-MeG) proceeds largely in an accurate but inefficient manner with a "wobble" base pairing between C and O(6)-MeG (Eoff, R. L., Irimia, A., Egli, M., and Guengerich, F. P. (2007) J. Biol. Chem. 282, 1456-1467). We considered here the bulky lesion O(6)-benzylguanine (O(6)-BzG) in DNA and catalysis by Dpo4. Mass spectrometry analysis of polymerization products revealed that the enzyme bypasses and extends across from O(6)-BzG, with C the major product ( approximately 70%) and some T and A ( approximately 15% each) incorporated opposite the lesion. Steady-state kinetic parameters indicated that Dpo4 was 7-, 5-, and 27-fold more efficient at C incorporation opposite O(6)-BzG than T, A, or G, respectively. In transient state kinetic analysis, the catalytic efficiency was decreased 62-fold for C incorporation opposite O(6)-BzG relative to unmodified DNA. Crystal structures reveal wobble pairing between C and O(6)-BzG. Pseudo-"Watson-Crick" pairing was observed between T and O(6)-BzG. Two other structures illustrate a possible mechanism for the accommodation of a +1 frameshift in the Dpo4 active site. The overall effect of O(6)-BzG is to decrease the efficiency of bypass by roughly an order of magnitude in every case except correct bypass, where the effect is not as pronounced. By comparison, Dpo4 is more accurate but no more efficient than model replicative polymerases, such as bacteriophage T7(-) DNA polymerase and human immunodeficiency virus-1 reverse transcriptase in the polymerization past O(6)-MeG and O(6)-BzG.

PubMed: 17337730
DOI: 10.1074/JBC.M700656200

Experimental method

X-RAY DIFFRACTION (2.39 Å)