TY - JOUR
T1 - Structure and mechanism of B-family DNA polymerase ζ specialized for translesion DNA synthesis
AU - Malik, Radhika
AU - Kopylov, Mykhailo
AU - Gomez-Llorente, Yacob
AU - Jain, Rinku
AU - Johnson, Robert E.
AU - Prakash, Louise
AU - Prakash, Satya
AU - Ubarretxena-Belandia, Iban
AU - Aggarwal, Aneel K.
N1 - Funding Information:
We thank B. Carragher, C. Potter and E. Eng for helpful advice and discussion throughout the project. We also thank Z. Zhang and D. Bobe for help in grid preparation, Y. Z. Tan for help in collecting tilted cryo-EM data, A. Brown and T. Terwilliger for help in implementing software and D. Nair for help in model building. This work was primarily funded by grant R01-GM124047 from the National Institutes of Health (NIH). I.U.-B. was supported by a grant PID2019-104423GB-I00/AEI/10.13039/501100011033 from the Spanish State Research Agency and by the Basque Excellence Research Centre program. Initial EM screening was performed at the Icahn School of Medicine microscope facility supported by a shared instrumentation grant from the NIH (1S10RR026473). Most of the cryo-EM work was performed at the Simons Electron Microscopy Center and National Resource for Automated Molecular Microscopy, located at the New York Structural Biology Center, supported by grants from the Simons Foundation (SF349247), NYSTAR and the NIH National Institute of General Medical Sciences (GM103310), with additional support from Agouron Institute (F00316), NIH (OD019994) and NIH (RR029300). Computing resources needed for this work were provided in part by the High Performance Computing facility of the Icahn School of Medicine at Mount Sinai. Molecular graphics and analyses were performed with UCSF Chimera, developed by the Resource for Biocomputing, Visualization, and Informatics at the University of California, San Francisco, with support from NIH P41-GM103311.
Publisher Copyright:
© 2020, The Author(s), under exclusive licence to Springer Nature America, Inc.
PY - 2020/10/1
Y1 - 2020/10/1
N2 - DNA polymerase ζ (Polζ) belongs to the same B-family as high-fidelity replicative polymerases, yet is specialized for the extension reaction in translesion DNA synthesis (TLS). Despite its importance in TLS, the structure of Polζ is unknown. We present cryo-EM structures of the Saccharomycescerevisiae Polζ holoenzyme in the act of DNA synthesis (3.1 Å) and without DNA (4.1 Å). Polζ displays a pentameric ring-like architecture, with catalytic Rev3, accessory Pol31‚ Pol32 and two Rev7 subunits forming an uninterrupted daisy chain of protein–protein interactions. We also uncover the features that impose high fidelity during the nucleotide-incorporation step and those that accommodate mismatches and lesions during the extension reaction. Collectively, we decrypt the molecular underpinnings of Polζ’s role in TLS and provide a framework for new cancer therapeutics.
AB - DNA polymerase ζ (Polζ) belongs to the same B-family as high-fidelity replicative polymerases, yet is specialized for the extension reaction in translesion DNA synthesis (TLS). Despite its importance in TLS, the structure of Polζ is unknown. We present cryo-EM structures of the Saccharomycescerevisiae Polζ holoenzyme in the act of DNA synthesis (3.1 Å) and without DNA (4.1 Å). Polζ displays a pentameric ring-like architecture, with catalytic Rev3, accessory Pol31‚ Pol32 and two Rev7 subunits forming an uninterrupted daisy chain of protein–protein interactions. We also uncover the features that impose high fidelity during the nucleotide-incorporation step and those that accommodate mismatches and lesions during the extension reaction. Collectively, we decrypt the molecular underpinnings of Polζ’s role in TLS and provide a framework for new cancer therapeutics.
UR - http://www.scopus.com/inward/record.url?scp=85089512881&partnerID=8YFLogxK
U2 - 10.1038/s41594-020-0476-7
DO - 10.1038/s41594-020-0476-7
M3 - Article
C2 - 32807989
AN - SCOPUS:85089512881
SN - 1545-9993
VL - 27
SP - 913
EP - 924
JO - Nature Structural and Molecular Biology
JF - Nature Structural and Molecular Biology
IS - 10
ER -