7m7s
Human DNA Pol eta S113A with dT-ended primer and 0.1 mM dAMPNPP
Structural highlights
DiseasePOLH_HUMAN Defects in POLH are the cause of xeroderma pigmentosum variant type (XPV) [MIM:278750; also designated as XP-V. Xeroderma pigmentosum (XP) is an autosomal recessive disease due to deficient nucleotide excision repair. It is characterized by hypersensitivity of the skin to sunlight, followed by high incidence of skin cancer and frequent neurologic abnormalities. XPV shows normal nucleotide excision repair, but an exaggerated delay in recovery of replicative DNA synthesis. Most XPV patients do not develop clinical symptoms and skin neoplasias until a later age. Clinical manifestations are limited to photo-induced deterioration of the skin and eyes.[1] [2] [3] [4] [5] FunctionPOLH_HUMAN DNA polymerase specifically involved in DNA repair. Plays an important role in translesion synthesis, where the normal high fidelity DNA polymerases cannot proceed and DNA synthesis stalls. Plays an important role in the repair of UV-induced pyrimidine dimers. Depending on the context, it inserts the correct base, but causes frequent base transitions and transversions. May play a role in hypermutation at immunoglobulin genes. Forms a Schiff base with 5'-deoxyribose phosphate at abasic sites, but does not have lyase activity. Targets POLI to replication foci.[6] [7] [8] [9] [10] Publication Abstract from PubMedDNA synthesis by polymerases is essential for life. Deprotonation of the nucleophile 3'-OH is thought to be the obligatory first step in the DNA synthesis reaction. We have examined each entity surrounding the nucleophile 3'-OH in the reaction catalyzed by human DNA polymerase (Pol) eta and delineated the deprotonation process by combining mutagenesis with steady-state kinetics, high-resolution structures of in crystallo reactions, and molecular dynamics simulations. The conserved S113 residue, which forms a hydrogen bond with the primer 3'-OH in the ground state, stabilizes the primer end in the active site. Mutation of S113 to alanine destabilizes primer binding and reduces the catalytic efficiency. Displacement of a water molecule that is hydrogen bonded to the 3'-OH using the 2'-OH of a ribonucleotide or 2'-F has little effect on catalysis. Moreover, combining the S113A mutation with 2'-F replacement, which removes two potential hydrogen acceptors of the 3'-OH, does not reduce the catalytic efficiency. We conclude that the proton can leave the O3' via alternative paths, supporting the hypothesis that binding of the third Mg(2+) initiates the reaction by breaking the alpha-beta phosphodiester bond of an incoming deoxyribonucleoside triphosphate (dNTP). Multiple deprotonation paths of the nucleophile 3'-OH in the DNA synthesis reaction.,Gregory MT, Gao Y, Cui Q, Yang W Proc Natl Acad Sci U S A. 2021 Jun 8;118(23):e2103990118. doi: , 10.1073/pnas.2103990118. PMID:34088846[11] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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