3UQ2

Crystal structure of the post-catalytic product complex of polymerase lambda with an rCMP inserted opposite a templating G and dAMP inserted opposite a templating T at the primer terminus.

Summary for 3UQ2

• Entry DOI: 10.2210/pdb3uq2/pdb
• Related: 3MGH 3MGI 3UPQ 3UQ0 3UQ1
• Descriptor: DNA polymerase lambda, 5'-D(*CP*GP*GP*CP*TP*GP*TP*AP*CP*TP*G)-3', 5'-D(*CP*AP*GP*TP*AP)-R(P*CP*A)-3', ... (8 entities in total)
• Functional Keywords: dna polymerase lambda, ribonucleotide incorporation, protein conformation, transferase, lyase-dna complex, lyase/dna
• Biological source: Homo sapiens (human)
• Cellular location: Nucleus: Q9UGP5
• Total number of polymer chains: 4
• Total formula weight: 43608.69

Authors

Gosavi, R.A.,Moon, A.F.,Kunkel, T.A.,Pedersen, L.C.,Bebenek, K. (deposition date: 2011-11-18, release date: 2012-05-23, Last modification date: 2023-09-13)

Primary citation

Gosavi, R.A.,Moon, A.F.,Kunkel, T.A.,Pedersen, L.C.,Bebenek, K.
The catalytic cycle for ribonucleotide incorporation by human DNA Pol lambda
Nucleic Acids Res., 40:7518-7527, 2012

PubMed Abstract: Although most DNA polymerases discriminate against ribonucleotide triphosphaets (rNTPs) during DNA synthesis, recent studies have shown that large numbers of ribonucleotides are incorporated into the eukaryotic nuclear genome. Here, we investigate how a DNA polymerase can stably incorporate an rNTP. The X-ray crystal structure of a variant of human DNA polymerase λ reveals that the rNTP occupies the nucleotide binding pocket without distortion of the active site, despite an unfavorable interaction between the 2'-O and Tyr505 backbone carbonyl. This indicates an energetically unstable binding state for the rNTP, stabilized by additional protein-nucleotide interactions. Supporting this idea is the 200-fold lower catalytic efficiency for rNTP relative to deoxyribonucleotide triphosphate (dNTP) incorporation, reflecting a higher apparent Km value for the rNTP. Furthermore, distortion observed in the structure of the post-catalytic product complex suggests that once the bond between the α- and β-phosphates of the rNTP is broken, the unfavorable binding state of the ribonucleotide cannot be maintained. Finally, structural and biochemical evaluation of dNTP insertion onto an ribonucleotide monophosphate (rNMP)-terminated primer indicates that a primer-terminal rNMP does not impede extension. The results are relevant to how ribonucleotides are incorporated into DNA in vivo, during replication and during repair, perhaps especially in non-proliferating cells when rNTP:dNTP ratios are high.

PubMed: 22584622
DOI: 10.1093/nar/gks413

Experimental method

X-RAY DIFFRACTION (2.25 Å)