As it has also been described for pol b, ASFV pol X displayed a limited stranddisplacement capacity, allowing further elongation of the primer strand.After incubation for min at C in the presence of ng of ASFV pol X, extension of the labeled strand was analyzed by electrophoresis in M urea, polyacrylamide gels and autoradiography.Despite the limited sensitivity of our in vitro misincorporation assay, it appears that fidelity of DNA synthesis by ASFV pol X relies on a proper selection of the complementary nucleotide as activator.It is conceivable that during evolution of DNA polymerases, a proper checking of the OH group of the nucleotide constituted an evolutionary advantage mainly for those polymerases involved in processive DNA synthesis, since the OH group of an incoming dNTP would be critical only for the next polymerization reaction, once dNMP is incorporated to the primer strand.A, scheme of the nucleotide gapped structure used as DNA substrate. B, top view of the palm and thumb subdomains of rat polb complexed with DNA, showing the amino acid residues that are invariant or highly conserved in ASFV pol X and the rest of pol X DNA polymerases.The palm subdomain, which consists largely of a bsheet, constitutes an appropriate surface to accommodate the substrate molecules and to present the pair of metal ions adequately for catalysis.The thumb subdomain forms a single wall of the original cleft described for the complete polbenzyme.In addition to these DNA binding core residues, resolution of the crystal structure of polbcomplexed with different DNA substrates Targetmol’s Captopril allowed the identification of two structurally similar, but functionally distinct, DNA binding motifs, named helixhairpinhelix binds the phosphate end of a DNA gap, allowing the removal of a deoxyribose phosphate intermediate prior to the gapfilling step during the BER reaction helps to position the primer strand of the DNA substrate in the purchase cysteine polymerase active site, and its coordinated metal ion was proposed to act as a processivity cofactor. These residues, always located in the palm subdomain, form a carboxylate triad implicated in the arrangement of a pair of divalent metal ions in a precise geometry to catalyze the nucleotidyl transfer reaction.As described for other DNA polymerases, mutation of these residues in polb. Much of the nucleotide binding site observed in the ternary complex of polb is made up of the terminus of the primer strand and the template overhang.Additionally, the two metal ions in the polbactive site bind to all three phosphates of the incoming nucleotide.It has also been noticed that closing of the thumb not only positions the dNTP, but also the templating base for an efficient and faithful nucleotide incorporation. It has also been proposed that a change back to the open conformation after nucleotide addition could be critical for facilitating the productoff step of catalysis, ratelimiting for the distributive mode of synthesis required for DNA repair. The conservation of the critical residues referred above and the fact that out of in vivo selected mutants of polbmap in the thumb subdomain, emphasizes its functional importance and suggests that a mobile thumb subdomain analogous to that of polbis operating also in the small ASFV pol X and the rest of polymerases from the pol X family.