Crystal structure of a thermostable type B DNA polymerase from Thermococcus gorgonarius.
Most known archaeal DNA polymerases belong to the type B family, which also includes the DNA replication polymerases of eukaryotes, but maintain high fidelity at extreme conditions. We describe here the 2.5 A resolution crystal structure of a DNA polymerase from the Archaea Thermococcus gorgonarius and identify structural features of the fold and the active site that are likely responsible for its thermostable function. Comparison with the mesophilic B type DNA polymerase gp43 of the bacteriophage RB69 highlights thermophilic adaptations, which include the presence of two disulfide bonds and an enhanced electrostatic complementarity at the DNA-protein interface. In contrast to gp43, several loops in the exonuclease and thumb domains are more closely packed; this apparently blocks primer binding to the exonuclease active site. A physiological role of this "closed" conformation is unknown but may represent a polymerase mode, in contrast to an editing mode with an open exonuclease site. This archaeal B DNA polymerase structure provides a starting point for structure-based design of polymerases or ligands with applications in biotechnology and the development of antiviral or anticancer agents.
Hopfner
K P
KP
Abteilung Strukturforschung, Max-Planck-Institut für Biochemie, D-82152 Martinsried, Germany. hopfner@scripps.edu
Eichinger
A
A
Engh
R A
RA
Laue
F
F
Ankenbauer
W
W
Huber
R
R
Angerer
B
B
D000595:Amino Acid Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D003196:Computer Graphics; D017124:Conserved Sequence; D018360:Crystallography, X-Ray / Q000379:methods; D004256:DNA Polymerase I / Q000737:chemistry* / Q000378:metabolism; D004795:Enzyme Stability; D006358:Hot Temperature; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D019712:Thermococcus / Q000201:enzymology*; D013816:Thermodynamics
D011994:Recombinant Proteins; D004256:DNA Polymerase I