An interaction network in the polymerase active site is a prerequisite for Watson-Crick base pairing in Pol γ

Park, Joon; Herrmann, Geoffrey K.; Roy, Arkanil; Shumate, Christie K.; Cisneros, G. Andrés; Yin, Y. Whitney

doi:10.1126/sciadv.adl3214

An interaction network in the polymerase active site is a prerequisite for Watson-Crick base pairing in Pol γ

Journal Article · Fri May 24 00:00:00 EDT 2024 · Science Advances

DOI:https://doi.org/10.1126/sciadv.adl3214· OSTI ID:2469664

^[1]; ^[2]; Roy, Arkanil ^[3]; Shumate, Christie K. ^[4]; ^[3]; ^[1]

University of Texas Medical Branch, Galveston, TX (United States)
University of Texas Medical Branch, Galveston, TX (United States); Baylor College of Medicine, Houston, TX (United States)
Univ. of Texas at Dallas, Richardson, TX (United States)
University of Texas Medical Branch, Galveston, TX (United States); University of Texas Health Science Center at Houston, TX (United States)

The replication accuracy of DNA polymerase gamma (Pol γ) is essential for mitochondrial genome integrity. Mutation of human Pol γ arginine-853 has been linked to neurological diseases. Although not a catalytic residue, Pol γ arginine-853 mutants are void of polymerase activity. To identify the structural basis for the disease, we determined a crystal structure of the Pol γ mutant ternary complex with correct incoming nucleotide 2'-deoxycytidine 5'-triphosphate (dCTP). Opposite to the wild type that undergoes open-to-closed conformational changes when bound to a correct nucleotide that is essential for forming a catalytically competent active site, the mutant complex failed to undergo the conformational change, and the dCTP did not base pair with its Watson-Crick complementary templating residue. Our studies revealed that arginine-853 coordinates an interaction network that aligns the 3'-end of primer and dCTP with the catalytic residues. Disruption of the network precludes the formation of Watson-Crick base pairing and closing of the active site, resulting in an inactive polymerase.

View Accepted Manuscript (DOE)

Research Organization:: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Basic Energy Sciences (BES); National Institutes of Health (NIH); National Science Foundation (NSF)

Grant/Contract Number:: AC02-05CH11231

OSTI ID:: 2469664

Journal Information:: Science Advances, Journal Name: Science Advances Journal Issue: 21 Vol. 10; ISSN 2375-2548

Publisher:: AAASCopyright Statement

Country of Publication:: United States

Language:: English

References (57)

Langevin dynamics of peptides: The frictional dependence of isomerization rates ofN-acetylalanyl-N?-methylamide Loncharich, Richard J.; Brooks, Bernard R.; Pastor, Richard W. Biopolymers, Vol. 32, Issue 5 https://doi.org/10.1002/bip.360320508	journal	May 1992
Molecular and clinical genetics of mitochondrial diseases due to POLG mutations Wong, Lee-Jun C.; Naviaux, Robert K.; Brunetti-Pierri, Nicola Human Mutation, Vol. 29, Issue 9 https://doi.org/10.1002/humu.20824	journal	September 2008
Development and testing of a general amber force field Wang, Junmei; Wolf, Romain M.; Caldwell, James W. Journal of Computational Chemistry, Vol. 25, Issue 9 https://doi.org/10.1002/jcc.20035	journal	January 2004
UCSF Chimera?A visualization system for exploratory research and analysis Pettersen, Eric F.; Goddard, Thomas D.; Huang, Conrad C. Journal of Computational Chemistry, Vol. 25, Issue 13 https://doi.org/10.1002/jcc.20084	journal	January 2004
Simplifying the clinical classification of polymerase gamma (POLG) disease based on age of onset; studies using a cohort of 155 cases Hikmat, Omar; Naess, Karin; Engvall, Martin Journal of Inherited Metabolic Disease, Vol. 43, Issue 4 https://doi.org/10.1002/jimd.12211	journal	January 2020
Mitochondrial mimicry of multiple system atrophy of the cerebellar subtype Mehta, Arpan R.; Fox, Susan H.; Tarnopolsky, Mark Movement Disorders, Vol. 26, Issue 4 https://doi.org/10.1002/mds.23510	journal	January 2011
MolProbity: More and better reference data for improved all-atom structure validation: PROTEIN SCIENCE.ORG Williams, Christopher J.; Headd, Jeffrey J.; Moriarty, Nigel W. Protein Science, Vol. 27, Issue 1 https://doi.org/10.1002/pro.3330	journal	November 2017
Current developments in Coot for macromolecular model building of Electron Cryo‐microscopy and Crystallographic Data Casañal, Ana; Lohkamp, Bernhard; Emsley, Paul Protein Science, Vol. 29, Issue 4 https://doi.org/10.1002/pro.3791	journal	March 2020
UCSF ChimeraX : Structure visualization for researchers, educators, and developers Pettersen, Eric F.; Goddard, Thomas D.; Huang, Conrad C. Protein Science, Vol. 30, Issue 1 https://doi.org/10.1002/pro.3943	journal	October 2020
Numerical integration of the cartesian equations of motion of a system with constraints: molecular dynamics of n-alkanes Ryckaert, Jean-Paul; Ciccotti, Giovanni; Berendsen, Herman J. C. Journal of Computational Physics, Vol. 23, Issue 3 https://doi.org/10.1016/0021-9991(77)90098-5	journal	March 1977
VMD: Visual molecular dynamics Humphrey, William; Dalke, Andrew; Schulten, Klaus Journal of Molecular Graphics, Vol. 14, Issue 1 https://doi.org/10.1016/0263-7855(96)00018-5	journal	February 1996
Side chains involved in catalysis of the polymerase reaction of DNA polymerase I from Escherichia coli. Polesky, A. H.; Dahlberg, M. E.; Benkovic, S. J. Journal of Biological Chemistry, Vol. 267, Issue 12 https://doi.org/10.1016/S0021-9258(18)42461-1	journal	April 1992
Identification of residues critical for the polymerase activity of the Klenow fragment of DNA polymerase I from Escherichia coli. Polesky, A. H.; Steitz, T. A.; Grindley, N. D. Journal of Biological Chemistry, Vol. 265, Issue 24 https://doi.org/10.1016/S0021-9258(18)77342-0	journal	August 1990
Protein Structure Prediction Using Rosetta Rohl, Carol A.; Strauss, Charlie E. M.; Misura, Kira M. S. Methods in Enzymology https://doi.org/10.1016/S0076-6879(04)83004-0	book	January 2004
[20] Processing of X-ray diffraction data collected in oscillation mode Otwinowski, Zbyszek; Minor, Wladek Macromolecular Crystallography Part A, 307-326 https://doi.org/10.1016/S0076-6879(97)76066-X	book	January 1997
Role of the mitochondrial DNA replication machinery in mitochondrial DNA mutagenesis, aging and age-related diseases DeBalsi, Karen L.; Hoff, Kirsten E.; Copeland, William C. Ageing Research Reviews, Vol. 33 https://doi.org/10.1016/j.arr.2016.04.006	journal	January 2017
Structural Insight into Processive Human Mitochondrial DNA Synthesis and Disease-Related Polymerase Mutations Lee, Young-Sam; Kennedy, W. Dexter; Yin, Y. Whitney Cell, Vol. 139, Issue 2 https://doi.org/10.1016/j.cell.2009.07.050	journal	October 2009
Directed Evolution of DNA Polymerase, RNA Polymerase and Reverse Transcriptase Activity in a Single Polypeptide Ong, Jennifer L.; Loakes, David; Jaroslawski, Szymon Journal of Molecular Biology, Vol. 361, Issue 3 https://doi.org/10.1016/j.jmb.2006.06.050	journal	August 2006
Fourier shell correlation threshold criteria van Heel, Marin; Schatz, Michael Journal of Structural Biology, Vol. 151, Issue 3 https://doi.org/10.1016/j.jsb.2005.05.009	journal	September 2005
One number does not fit all: Mapping local variations in resolution in cryo-EM reconstructions Cardone, Giovanni; Heymann, J. Bernard; Steven, Alasdair C. Journal of Structural Biology, Vol. 184, Issue 2 https://doi.org/10.1016/j.jsb.2013.08.002	journal	November 2013
Characterization of Nine Cancer-Associated Variants in Human DNA Polymerase κ Antczak, Nicole M.; Walker, Alice R.; Stern, Hannah R. Chemical Research in Toxicology, Vol. 31, Issue 8 https://doi.org/10.1021/acs.chemrestox.8b00055	journal	July 2018
ff14SB: Improving the Accuracy of Protein Side Chain and Backbone Parameters from ff99SB Maier, James A.; Martinez, Carmenza; Kasavajhala, Koushik Journal of Chemical Theory and Computation, Vol. 11, Issue 8 https://doi.org/10.1021/acs.jctc.5b00255	journal	July 2015
Assessing the Current State of Amber Force Field Modifications for DNA Galindo-Murillo, Rodrigo; Robertson, James C.; Zgarbová, Marie Journal of Chemical Theory and Computation, Vol. 12, Issue 8 https://doi.org/10.1021/acs.jctc.6b00186	journal	July 2016
Computational Characterization of the Inhibition Mechanism of Xanthine Oxidoreductase by Topiroxostat Maghsoud, Yazdan; Dong, Chao; Cisneros, G. Andrés ACS Catalysis, Vol. 13, Issue 9 https://doi.org/10.1021/acscatal.3c01245	journal	April 2023
Correlation of the Kinetics of Finger Domain Mutants in RB69 DNA Polymerase with Its Structure ^† Yang, Guangwei; Franklin, Matthew; Li, Jing Biochemistry, Vol. 41, Issue 8 https://doi.org/10.1021/bi0119924	journal	February 2002
Characterization of the Native and Recombinant Catalytic Subunit of Human DNA Polymerase γ: Identification of Residues Critical for Exonuclease Activity and Dideoxynucleotide Sensitivity Longley, Matthew J.; Ropp, Philip A.; Lim, Susan E. Biochemistry, Vol. 37, Issue 29 https://doi.org/10.1021/bi980772w	journal	July 1998
Human Mitochondrial DNA Polymerase Holoenzyme: Reconstitution and Characterization Johnson, Allison A.; Tsai, Yu-chih; Graves, Steven W. Biochemistry, Vol. 39, Issue 7 https://doi.org/10.1021/bi992104w	journal	January 2000
PROPKA3: Consistent Treatment of Internal and Surface Residues in Empirical p K _a Predictions Olsson, Mats H. M.; Søndergaard, Chresten R.; Rostkowski, Michal Journal of Chemical Theory and Computation, Vol. 7, Issue 2 https://doi.org/10.1021/ct100578z	journal	December 2010
PTRAJ and CPPTRAJ: Software for Processing and Analysis of Molecular Dynamics Trajectory Data Roe, Daniel R.; Cheatham, Thomas E. Journal of Chemical Theory and Computation, Vol. 9, Issue 7 https://doi.org/10.1021/ct400341p	journal	June 2013
Insight into Enzymatic C−F Bond Formation from QM and QM/MM Calculations Senn, Hans Martin; O'Hagan, David; Thiel, Walter Journal of the American Chemical Society, Vol. 127, Issue 39 https://doi.org/10.1021/ja053875s	journal	September 2005
Distal Mutations in the β-Clamp of DNA Polymerase III* Disrupt DNA Orientation and Affect Exonuclease Activity Berger, Madison B.; Cisneros, G. Andrés Journal of the American Chemical Society, Vol. 145, Issue 6 https://doi.org/10.1021/jacs.2c11713	journal	February 2023
The fidelity of DNA synthesis by eukaryotic replicative and translesion synthesis polymerases McCulloch, Scott D.; Kunkel, Thomas A. Cell Research, Vol. 18, Issue 1 https://doi.org/10.1038/cr.2008.4	journal	January 2008
cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination Punjani, Ali; Rubinstein, John L.; Fleet, David J. Nature Methods, Vol. 14, Issue 3 https://doi.org/10.1038/nmeth.4169	journal	February 2017
Integration and global analysis of isothermal titration calorimetry data for studying macromolecular interactions Brautigam, Chad A.; Zhao, Huaying; Vargas, Carolyn Nature Protocols, Vol. 11, Issue 5 https://doi.org/10.1038/nprot.2016.044	journal	April 2016
Local computational methods to improve the interpretability and analysis of cryo-EM maps Kaur, Satinder; Gomez-Blanco, Josue; Khalifa, Ahmad A. Z. Nature Communications, Vol. 12, Issue 1 https://doi.org/10.1038/s41467-021-21509-5	journal	February 2021
Non-uniform refinement: adaptive regularization improves single-particle cryo-EM reconstruction Punjani, Ali; Zhang, Haowei; Fleet, David J. Nature Methods, Vol. 17, Issue 12 https://doi.org/10.1038/s41592-020-00990-8	journal	November 2020
Measurement of atom resolvability in cryo-EM maps with Q-scores Pintilie, Grigore; Zhang, Kaiming; Su, Zhaoming Nature Methods, Vol. 17, Issue 3 https://doi.org/10.1038/s41592-020-0731-1	journal	February 2020
Polγ coordinates DNA synthesis and proofreading to ensure mitochondrial genome integrity Park, Joon; Herrmann, Geoffrey K.; Mitchell, Patrick G. Nature Structural & Molecular Biology, Vol. 30, Issue 6 https://doi.org/10.1038/s41594-023-00980-2	journal	May 2023
Alpha-carboxy nucleoside phosphonates as universal nucleoside triphosphate mimics Balzarini, Jan; Das, Kalyan; Bernatchez, Jean A. Proceedings of the National Academy of Sciences, Vol. 112, Issue 11 https://doi.org/10.1073/pnas.1420233112	journal	March 2015
Comparison between DNA melting thermodynamics and DNA polymerase fidelity. Petruska, J.; Goodman, M. F.; Boosalis, M. S. Proceedings of the National Academy of Sciences, Vol. 85, Issue 17 https://doi.org/10.1073/pnas.85.17.6252	journal	September 1988
Side Chains That Influence Fidelity at the Polymerase Active Site of Escherichia coli DNA Polymerase I (Klenow Fragment) Minnick, Dana T.; Bebenek, Katarzyna; Osheroff, Wendy P. Journal of Biological Chemistry, Vol. 274, Issue 5 https://doi.org/10.1074/jbc.274.5.3067	journal	January 1999
Characterization of a Catalytically Slow AP Lyase Activity in DNA Polymerase γ and Other Family A DNA Polymerases Pinz, Kevin G.; Bogenhagen, Daniel F. Journal of Biological Chemistry, Vol. 275, Issue 17 https://doi.org/10.1074/jbc.275.17.12509	journal	April 2000
Escherichia coli DNA Polymerase I (Klenow Fragment) Uses a Hydrogen-bonding Fork from Arg668 to the Primer Terminus and Incoming Deoxynucleotide Triphosphate to Catalyze DNA Replication Meyer, Aviva S.; Blandino, Maureen; Spratt, Thomas E. Journal of Biological Chemistry, Vol. 279, Issue 32 https://doi.org/10.1074/jbc.C400232200	journal	August 2004
Disease Mutations in the Human Mitochondrial DNA Polymerase Thumb Subdomain Impart Severe Defects in Mitochondrial DNA Replication Kasiviswanathan, Rajesh; Longley, Matthew J.; Chan, Sherine S. L. Journal of Biological Chemistry, Vol. 284, Issue 29 https://doi.org/10.1074/jbc.M109.011940	journal	July 2009
The Properties of Steric Gate Mutants Reveal Different Constraints within the Active Sites of Y-family and A-family DNA Polymerases DeLucia, Angela M.; Chaudhuri, Santanov; Potapova, Olga Journal of Biological Chemistry, Vol. 281, Issue 37 https://doi.org/10.1074/jbc.M604393200	journal	July 2006
ProDy 2.0: increased scale and scope after 10 years of protein dynamics modelling with Python Zhang, She; Krieger, James M.; Zhang, Yan Bioinformatics, Vol. 37, Issue 20 https://doi.org/10.1093/bioinformatics/btab187	journal	April 2021
Accuracy, lesion bypass, strand displacement and translocation by DNA polymerases Steitz, Thomas A.; Yin, Y. Whitney Philosophical Transactions of the Royal Society of London. Series B: Biological Sciences, Vol. 359, Issue 1441 https://doi.org/10.1098/rstb.2003.1374	journal	January 2004
Coot model-building tools for molecular graphics Emsley, Paul; Cowtan, Kevin Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 12, p. 2126-2132 https://doi.org/10.1107/S0907444904019158	journal	November 2004
Estimation of high-order aberrations and anisotropic magnification from cryo-EM data sets in RELION -3.1 Zivanov, Jasenko; Nakane, Takanori; Scheres, Sjors H. W. IUCrJ, Vol. 7, Issue 2 https://doi.org/10.1107/S2052252520000081	journal	February 2020
ISOLDE : a physically realistic environment for model building into low-resolution electron-density maps Croll, Tristan Ian Acta Crystallographica Section D Structural Biology, Vol. 74, Issue 6 https://doi.org/10.1107/S2059798318002425	journal	April 2018
Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix Liebschner, Dorothee; Afonine, Pavel V.; Baker, Matthew L. Acta Crystallographica Section D Structural Biology, Vol. 75, Issue 10 https://doi.org/10.1107/S2059798319011471	journal	October 2019
Matplotlib: A 2D Graphics Environment Hunter, John D. Computing in Science & Engineering, Vol. 9, Issue 3 https://doi.org/10.1109/MCSE.2007.55	journal	January 2007
Mitochondrial DNA polymerase mutations: an ever expanding molecular and clinical spectrum Tang, S.; Wang, J.; Lee, N. -C. Journal of Medical Genetics, Vol. 48, Issue 10 https://doi.org/10.1136/jmedgenet-2011-100222	journal	August 2011
Novel POLG1 mutations associated with neuromuscular and liver phenotypes in adults and children Stewart, J. D.; Tennant, S.; Powell, H. Journal of Medical Genetics, Vol. 46, Issue 3 https://doi.org/10.1136/jmg.2008.058180	journal	March 2008
DNA Replication Fidelity Kunkel, Thomas A.; Bebenek, Katarzyna Annual Review of Biochemistry, Vol. 69, Issue 1 https://doi.org/10.1146/annurev.biochem.69.1.497	journal	June 2000
Structural basis for processivity and antiviral drug toxicity in human mitochondrial DNA replicase Szymanski, Michal R.; Kuznetsov, Vladmir B.; Shumate, Christie The EMBO Journal, Vol. 34, Issue 14 https://doi.org/10.15252/embj.201591520	journal	June 2015
Biochemical Basis of DNA Replication Fidelity Goodman, Myron F.; Creighton, Steven; Bloom, Linda B. Critical Reviews in Biochemistry and Molecular Biology, Vol. 28, Issue 2 https://doi.org/10.3109/10409239309086792	journal	January 1993

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An interaction network in the polymerase active site is a prerequisite for Watson-Crick base pairing in Pol γ

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