Top view of the crystal structure of the replicative DNA polymerase DnaE1 from Mycobacterium tuberculosis (Mtb). Mtb DnaE1 uses an intrinsic exonuclease located in the PHP domain of the polymerase (shown in red). The PHP-exonuclease has nine conserved residues that coordinate a trinuclear zinc center, contrary to almost all high-fidelity DNA polymerases that make use of magnesium-dependent exonucleases.
Baños-Mateos S, van Roon AM, Lang UF, Maslen SL, Skehel JM, Lamers MH.
Nat Commun2017 Oct; 8: 855.
High-fidelity DNA replication depends on a proofreading 3′-5′ exonuclease that is associated with the replicative DNA polymerase. The replicative DNA polymerase DnaE1 from the major pathogen Mycobacterium tuberculosis (Mtb) uses its intrinsic PHP-exonuclease that is distinct from the canonical DEDD exonucleases found in the Escherichia coli and eukaryotic replisomes. The mechanism of the PHP-exonuclease is not known. Here, we present the crystal structure of the Mtb DnaE1 polymerase. The PHP-exonuclease has a trinuclear zinc center, coordinated by nine conserved residues. Cryo-EM analysis reveals the entry path of the primer strand in the PHP-exonuclease active site. Furthermore, the PHP-exonuclease shows a striking similarity to E. coli endonuclease IV, which provides clues regarding the mechanism of action. Altogether, this work provides important insights into the PHP-exonuclease and reveals unique properties that make it an attractive target for novel anti-mycobacterial drugs.The polymerase and histidinol phosphatase (PHP) domain in the DNA polymerase DnaE1 is essential for mycobacterial high-fidelity DNA replication. Here, the authors determine the DnaE1 crystal structure, which reveals the PHP-exonuclease mechanism that can be exploited for antibiotic development.