Dpo4 is hindered in extending a G.T mismatch by a reverse wobble.
The ability or inability of a DNA polymerase to extend a mispair directly affects the establishment of genomic mutations. We report here kinetic analyses of the ability of Dpo4, a Y-family polymerase from Sulfolobus solfataricus, to extend from all mispairs opposite a template G or T. Dpo4 is equally inefficient at extending these mispairs, which include, surprisingly, a G.T mispair expected to conform closely to Watson-Crick geometry. To elucidate the basis of this, we solved the structure of Dpo4 bound to G.T-mispaired primer template in the presence of an incoming nucleotide. As a control, we also determined the structure of Dpo4 bound to a matched A-T base pair at the primer terminus. The structures offer a basis for the low efficiency of Dpo4 in extending a G.T mispair: a reverse wobble that deflects the primer 3'-OH away from the incoming nucleotide.
Trincao
Jose
J
Structural Biology Program, Department of Physiology and Biophysics, Mount Sinai School of Medicine, Box 1677, 1425 Madison Avenue, New York, New York 10029, USA.
Johnson
Robert E
RE
Wolfle
William T
WT
Escalante
Carlos R
CR
Prakash
Satya
S
Prakash
Louise
L
Aggarwal
Aneel K
AK
D001426:Bacterial Proteins / Q000737:chemistry*; D020137:Base Pair Mismatch; D001483:Base Sequence; D018360:Crystallography, X-Ray; D017931:DNA Primers; D007700:Kinetics; D008958:Models, Molecular; D011487:Protein Conformation; D016944:Sulfolobus / Q000201:enzymology
D001426:Bacterial Proteins; D017931:DNA Primers