9EA2

T4 Bacteriophage Replicative Polymerase Captured in Polymerase Exchange State 2

Summary for 9EA2

• Entry DOI: 10.2210/pdb9ea2/pdb
• EMDB information: 47824
• Descriptor: DNA-directed DNA polymerase, Sliding clamp, DNA (5'-AGC TAT GAC CAT GAT TAC GAA TTG ddC-3'), ... (4 entities in total)
• Functional Keywords: replication, t4, holoenzyme, replication-dna complex, replication/dna
• Biological source: Escherichia phage T4; More
• Total number of polymer chains: 7
• Total formula weight: 301408.79

Authors

Li, H.,Feng, X. (deposition date: 2024-11-10, release date: 2025-11-19, Last modification date: 2026-01-14)

Primary citation

Feng, X.,Spiering, M.M.,Ruda de Luna Santos, A.,Benkovic, S.J.,Li, H.
Structural insights into the exchange mechanism of a replicative DNA polymerase.
Nucleic Acids Res., 53:-, 2025

PubMed Abstract: Replicative DNA polymerases are distinguished by their speed, processivity, and fidelity. While speed and fidelity arise from the polymerase's intrinsic catalytic and proofreading activities, processivity is typically attributed to the DNA sliding clamp that tethers the polymerase to DNA. However, additional mechanisms may also contribute. The T4 bacteriophage polymerase can exchange on-the-fly, a process likely contributing to its ∼10-fold higher synthesis rate compared with human polymerases. Here, we reconstituted the T4 holoenzyme and polymerase exchange complexes using purified gp43 polymerase, gp45 sliding clamp, and a primer-template DNA substrate. Cryo-electron microscopy (cryo-EM) analysis revealed either one or two polymerases bound to the clamp and DNA. In the one-polymerase complex, the DNA threads perpendicularly through the clamp, supporting processive synthesis. In contrast, the two-polymerase complex displays a markedly tilted DNA orientation, impeding sliding and representing exchange intermediates. Three distinct conformational states of the two-polymerase complex define a multistep exchange mechanism. To our knowledge, these findings provide the first molecular-level view of replicative polymerase exchange.

PubMed: 41459747
DOI: 10.1093/nar/gkaf1359

Experimental method

ELECTRON MICROSCOPY (3.7 Å)