6GO6

TdT chimera (Loop1 of pol mu) - ternary complex with downstream dsDNA

Summary for 6GO6

• Entry DOI: 10.2210/pdb6go6/pdb
• Descriptor: DNA nucleotidylexotransferase,DNA-directed DNA/RNA polymerase mu,DNA nucleotidylexotransferase, DNA (5'-D(*TP*TP*TP*TP*TP*GP*GP*C)-3'), DNA (5'-D(*AP*AP*AP*AP*AP*C)-3'), ... (7 entities in total)
• Functional Keywords: nhej pathway, dna bridging, dna polymerase polx, dna binding protein
• Biological source: Mus musculus (Mouse); More
• Total number of polymer chains: 4
• Total formula weight: 53004.89

Authors

Loc'h, J.,Gerodimos, C.A.,Rosario, S.,Lieber, M.R.,Delarue, M. (deposition date: 2018-06-01, release date: 2019-06-05, Last modification date: 2024-01-17)

Primary citation

Loc'h, J.,Gerodimos, C.A.,Rosario, S.,Tekpinar, M.,Lieber, M.R.,Delarue, M.
Structural evidence for an intransbase selection mechanism involving Loop1 in polymerase mu at an NHEJ double-strand break junction.
J.Biol.Chem., 294:10579-10595, 2019

PubMed Abstract: Eukaryotic DNA polymerase (Pol) X family members such as Pol μ and terminal deoxynucleotidyl transferase (TdT) are important components for the nonhomologous DNA end-joining (NHEJ) pathway. TdT participates in a specialized version of NHEJ, V(D)J recombination. It has primarily nontemplated polymerase activity but can take instructions across strands from the downstream dsDNA, and both activities are highly dependent on a structural element called Loop1. However, it is unclear whether Pol μ follows the same mechanism, because the structure of its Loop1 is disordered in available structures. Here, we used a chimeric TdT harboring Loop1 of Pol μ that recapitulated the functional properties of Pol μ in ligation experiments. We solved three crystal structures of this TdT chimera bound to several DNA substrates at 1.96-2.55 Å resolutions, including a full DNA double-strand break (DSB) synapsis. We then modeled the full Pol μ sequence in the context of one these complexes. The atomic structure of an NHEJ junction with a Pol X construct that mimics Pol μ in a reconstituted system explained the distinctive properties of Pol μ compared with TdT. The structure suggested a mechanism of base selection relying on Loop1 and taking instructions via the in templating base independently of the primer strand. We conclude that our atomic-level structural observations represent a paradigm shift for the mechanism of base selection in the Pol X family of DNA polymerases.

PubMed: 31138645
DOI: 10.1074/jbc.RA119.008739

Experimental method

X-RAY DIFFRACTION (2.09 Å)