2j6t: Difference between revisions

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Revision as of 16:24, 23 March 2008

File:2j6t.gif

, resolution 2.60Å

Sites:

Ligands:

and

Activity:

DNA-directed DNA polymerase, with EC number 2.7.7.7

Coordinates:

save as pdb, mmCIF, xml

TERNARY COMPLEX OF SULFOLOBUS SOLFATARICUS DPO4 DNA POLYMERASE, O6-METHYLGUANINE MODIFIED DNA, AND DATP.

OverviewOverview

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.

About this StructureAbout this Structure

2J6T is a Single protein structure of sequence from Sulfolobus solfataricus. Full crystallographic information is available from OCA.

ReferenceReference

Sulfolobus solfataricus DNA polymerase Dpo4 is partially inhibited by "wobble" pairing between O6-methylguanine and cytosine, but accurate bypass is preferred., Eoff RL, Irimia A, Egli M, Guengerich FP, J Biol Chem. 2007 Jan 12;282(2):1456-67. Epub 2006 Nov 14. PMID:17105728

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OCA

@@ Line 4: / Line 4: @@
 |PDB= 2j6t |SIZE=350|CAPTION= <scene name='initialview01'>2j6t</scene>, resolution 2.60&Aring;
 |SITE= <scene name='pdbsite=AC1:Ca+Binding+Site+For+Chain+A'>AC1</scene>
-|LIGAND= <scene name='pdbligand=CA:CALCIUM+ION'>CA</scene> and <scene name='pdbligand=DTP:2'-DEOXYADENOSINE 5'-TRIPHOSPHATE'>DTP</scene>
+|LIGAND= <scene name='pdbligand=CA:CALCIUM+ION'>CA</scene> and <scene name='pdbligand=DTP:2&#39;-DEOXYADENOSINE 5&#39;-TRIPHOSPHATE'>DTP</scene>
 |ACTIVITY= [http://en.wikipedia.org/wiki/DNA-directed_DNA_polymerase DNA-directed DNA polymerase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.7.7.7 2.7.7.7]
 |GENE=
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 [[Category: translesion dna synthesis]]
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Mar 20 17:38:03 2008''
+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Mar 23 15:24:36 2008''