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Structural basis for the binding and incorporation of nucleotide analogs with L-stereochemistry by human DNA polymerase λ.

Vyas R, Zahurancik WJ, Suo Z
Proc Natl Acad Sci U S A 111 E3033-42 (2014)
Related entries: 4k4g, 4k4h

Cited: 8 times
EuropePMC logo PMID: 25015085

Abstract

Although lamivudine and emtricitabine, two L-deoxycytidine analogs, have been widely used as antiviral drugs for years, a structural basis for D-stereoselectivity against L-dNTPs, enantiomers of natural nucleotides (D-dNTPs), by any DNA polymerase or reverse transcriptase has not been established due to lack of a ternary structure of a polymerase, DNA, and an incoming L-dNTP. Here, we report 2.10-2.25 Å ternary crystal structures of human DNA polymerase λ, DNA, and L-deoxycytidine 5'-triphosphate (L-dCTP), or the triphosphates of lamivudine ((-)3TC-TP) and emtricitabine ((-)FTC-TP) with four ternary complexes per asymmetric unit. The structures of these 12 ternary complexes reveal that relative to D-deoxycytidine 5'-triphosphate (D-dCTP) in the canonical ternary structure of Polλ-DNA-D-dCTP, L-dCTP, (-)3TC-TP, and (-)FTC-TP all have their ribose rotated by 180°. Among the four ternary complexes with a specific L-nucleotide, two are similar and show that the L-nucleotide forms three Watson-Crick hydrogen bonds with the templating nucleotide dG and adopts a chair-like triphosphate conformation. In the remaining two similar ternary complexes, the L-nucleotide surprisingly interacts with the side chain of a conserved active site residue R517 through one or two hydrogen bonds, whereas the templating dG is anchored by a hydrogen bond with the side chain of a semiconserved residue Y505. Furthermore, the triphosphate of the L-nucleotide adopts an unprecedented N-shaped conformation. Our mutagenic and kinetic studies further demonstrate that the side chain of R517 is critical for the formation of the abovementioned four complexes along proposed catalytic pathways for L-nucleotide incorporation and provide the structural basis for the D-stereoselectivity of a DNA polymerase.

Articles - 4k4i mentioned but not cited (2)

  1. Elucidating molecular interactions of L-nucleotides with HIV-1 reverse transcriptase and mechanism of M184V-caused drug resistance. Hung M, Tokarsky EJ, Lagpacan L, Zhang L, Suo Z, Lansdon EB. Commun Biol 2 469 (2019)
  2. Structural basis for the binding and incorporation of nucleotide analogs with L-stereochemistry by human DNA polymerase λ. Vyas R, Zahurancik WJ, Suo Z. Proc Natl Acad Sci U S A 111 E3033-42 (2014)


Articles citing this publication (6)

  1. Structural insights into the recognition of nucleoside reverse transcriptase inhibitors by HIV-1 reverse transcriptase: First crystal structures with reverse transcriptase and the active triphosphate forms of lamivudine and emtricitabine. Bertoletti N, Chan AH, Schinazi RF, Yin YW, Anderson KS. Protein Sci 28 1664-1675 (2019)
  2. Significant impact of divalent metal ions on the fidelity, sugar selectivity, and drug incorporation efficiency of human PrimPol. Tokarsky EJ, Wallenmeyer PC, Phi KK, Suo Z. DNA Repair (Amst) 49 51-59 (2017)
  3. Structural basis for the D-stereoselectivity of human DNA polymerase β. Vyas R, Reed AJ, Raper AT, Zahurancik WJ, Wallenmeyer PC, Suo Z. Nucleic Acids Res 45 6228-6237 (2017)
  4. Digital gene expression profiling analysis of DNA repair pathways in colon cancer stem population of HT29 cells. Wang W, Zhang G, Yang J, Gu H, Ding L, Yu H, Yu M, Cui Q, Ji X, Li M. Acta Biochim Biophys Sin (Shanghai) 49 90-100 (2017)
  5. Compatibility and Fidelity of Mirror-Image Thymidine in Transcription Events by T7 RNA Polymerase. Liu Q, Ke Y, Kan Y, Tang X, Li X, He Y, Wu L. Mol Ther Nucleic Acids 21 604-613 (2020)
  6. Advances in Structural and Single-Molecule Methods for Investigating DNA Lesion Bypass and Repair Polymerases. Raper AT, Reed AJ, Gadkari VV, Suo Z. Chem Res Toxicol 30 260-269 (2017)


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