2VWJ: Uracil Recognition In Archaeal Dna Polymerases Captured By X-Ray Crystallography

Citation:
Abstract
Archaeal family B DNA polymerases bind tightly to template-strand uracil and stall replication on encountering the pro-mutagenic base. This article describes an X-ray crystal structure, at 2.8 A resolution, of Thermococcus gorgonarius polymerase in complex with a DNA primer-template containing uracil in the single-stranded region. The DNA backbone is distorted to position the uracil deeply within a pocket, located in the amino-terminal domain of the polymerase. Specificity arises from a combination of hydrogen bonds between the protein backbone and uracil, with the pocket shaped to prevent the stable binding of the four standard DNA bases. Strong interactions are seen with the two phosphates that flank the uracil and the structure gives clues concerning the coupling of uracil binding to the halting of replication. The importance of key amino acids, identified by the analysis of the structure and their conservation between archaeal polymerases, was confirmed by site-directed mutagenesis. The crystal structure of V93Q, a polymerase variant that no longer recognises uracil, is also reported, explaining the V93Q phenotype by the steric exclusion of uracil from the pocket.
PDB ID: 2VWJDownload
MMDB ID: 65590
PDB Deposition Date: 2008/6/25
Updated in MMDB: 2012/11
Experimental Method:
x-ray diffraction
Resolution: 2.78  Å
Source Organism:
Thermococcus gorgonarius
Similar Structures:
Biological Unit for 2VWJ: dimeric; determined by author and by software (PISA)
Molecular Components in 2VWJ
Label Count Molecule
Protein (1 molecule)
1
DNA Polymerase
Molecule annotation
Nucleotide(1 molecule)
1
5'-d(*ap*ap*up*gp*gp*ap*gp*ap*cp*gp *gp*cp*tp*tp*tp*tp*gp*cp*cp*gp*tp*gp*tp*c)-3'
Molecule annotation
Chemicals (3 molecules)
1
3
* Click molecule labels to explore molecular sequence information.

Citing MMDB
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