Open-system thermodynamic analysis of DNA polymerase fidelity

Replication inside a living cell, carried out by DNA polymerase, has an error rate far below that predicted by equilibrium thermodynamics from the affinities between nucleotides and a polymerase complex. The high fidelity is achieved through several distinctly different molecular mechanisms that include a nucleotide insertion checkpoint and 3'-5' exonuclease activity. The checkpoint mechanism has recently been articulated as a new paradigm for high specificity. A rigorous thermodynamic analysis of the bare DNA polymerization reaction, i.e., in the absence of exonuclease activity and proofreading, is developed in this paper. The analysis (a) reveals the important role of nonequilibrium steady-state (NESS) flux that drives high fidelity, (b) quantifies the error rate of the polymerization reaction as a function of free energy input through sustained non-equilibrium between chemical species, (c) bridges the 'thermodynamic' and 'kinetic' views of specificity and (d) generalizes the theory of kinetic checkpoints and provides it with a sound thermodynamic basis. The underlying mechanism again calls attention to the energy expenditure in heightened biomolecular specificity, a concept first developed by Hopfield and Ninio in the mid-1970s. The mechanism discussed in the present paper is not limited to DNA replication alone; it may be applicable to other biochemical systems.

[1]  D. Fygenson,et al.  DNA polymerase fidelity: from genetics toward a biochemical understanding. , 1998, Genetics.

[2]  William S. Hlavacek,et al.  Kinetic proofreading models for cell signaling predict ways to escape kinetic proofreading , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[3]  J. Durup.  On the relations between error rates in DNA replication and elementary chemical rate constants. , 1982, Journal of theoretical biology.

[4]  Hong Qian,et al.  Open-system nonequilibrium steady state: statistical thermodynamics, fluctuations, and chemical oscillations. , 2006, The journal of physical chemistry. B.

[5]  S. Benkovic,et al.  DNA polymerase fidelity: kinetics, structure, and checkpoints , 2004 .

[6]  J. Heinonen.  Biological Role of Inorganic Pyrophosphate , 2001, Springer US.

[7]  T. L. Hill,et al.  Free Energy Transduction and Biochemical Cycle Kinetics , 1988, Springer New York.

[8]  J. Puglisi,et al.  tRNA selection and kinetic proofreading in translation , 2004, Nature Structural &Molecular Biology.

[9]  Robert A Beckman,et al.  Genetic instability in cancer: theory and experiment. , 2005, Seminars in cancer biology.

[10]  Hong Qian,et al.  Reducing intrinsic biochemical noise in cells and its thermodynamic limit. , 2006, Journal of molecular biology.

[11]  B. Stillman,et al.  Constitutively high dNTP concentration inhibits cell cycle progression and the DNA damage checkpoint in yeast Saccharomyces cerevisiae , 2007, Proceedings of the National Academy of Sciences.

[12]  P. V. von Hippel,et al.  The elongation-termination decision in transcription. , 1992, Science.

[13]  Are further kinetic amplification schemes possible? , 1977, Biochimie.

[14]  T. Kunkel,et al.  Fidelity of DNA synthesis. , 1982, Annual review of biochemistry.

[15]  Guobin Luo,et al.  Single-molecule and ensemble fluorescence assays for a functionally important conformational change in T7 DNA polymerase , 2007, Proceedings of the National Academy of Sciences.

[16]  H. Fearnhead,et al.  Intracellular Nucleotides Act as Critical Prosurvival Factors by Binding to Cytochrome C and Inhibiting Apoptosome , 2006, Cell.

[17]  K. Breslauer,et al.  The thermodynamics of template-directed DNA synthesis: Base insertion and extension enthalpies , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[18]  M. Goodman,et al.  Hydrogen bonding revisited: geometric selection as a principal determinant of DNA replication fidelity. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[19]  H. Qian.  Cooperativity and specificity in enzyme kinetics: a single-molecule time-based perspective. , 2008, Biophysical journal.

[20]  J. Hopfield.  The energy relay: a proofreading scheme based on dynamic cooperativity and lacking all characteristic symptoms of kinetic proofreading in DNA replication and protein synthesis. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[21]  Hong Qian,et al.  Generalized Haldane equation and fluctuation theorem in the steady-state cycle kinetics of single enzymes. , 2005, Physical review. E, Statistical, nonlinear, and soft matter physics.

[22]  J. Hopfield.  Kinetic proofreading: a new mechanism for reducing errors in biosynthetic processes requiring high specificity. , 1974, Proceedings of the National Academy of Sciences of the United States of America.

[23]  D. Galas,et al.  Enzymatic determinants of DNA polymerase accuracy. Theory of coliphage T4 polymerase mechanisms. , 1978, Journal of molecular biology.

[24]  P. Swain,et al.  The role of proofreading in signal transduction specificity. , 2002, Biophysical journal.

[25]  Erin K O'Shea,et al.  Partially Phosphorylated Pho4 Activates Transcription of a Subset of Phosphate-Responsive Genes , 2003, PLoS biology.

[26]  M. Fisher,et al.  Molecular motors: a theorist's perspective. , 2007, Annual review of physical chemistry.

[27]  M. Magnasco,et al.  A kinetic proofreading mechanism for disentanglement of DNA by topoisomerases , 1999, Nature.

[28]  Q Zheng,et al.  Progress of a half century in the study of the Luria-Delbrück distribution. , 1999, Mathematical biosciences.

[29]  T. Kunkel,et al.  On the fidelity of DNA synthesis. Pyrophosphate-induced misincorporation allows detection of two proofreading mechanisms. , 1986, Journal of Biological Chemistry.

[30]  J. Ninio.  Kinetic amplification of enzyme discrimination. , 1975, Biochimie.

[31]  Yuhai Tu,et al.  The nonequilibrium mechanism for ultrasensitivity in a biological switch: Sensing by Maxwell's demons , 2008, Proceedings of the National Academy of Sciences.

[32]  A. Kornberg,et al.  Enzymatic Synthesis of Deoxyribonucleic Acid XXXVI. A PROOFREADING FUNCTION FOR THE 3' → 5' EXONUCLEASE ACTIVITY IN DEOXYRIBONUCLEIC ACID POLYMERASES , 1972 .

[33]  Richard Wolfenden,et al.  Thermodynamic and extrathermodynamic requirements of enzyme catalysis. , 2003, Biophysical chemistry.

[34]  Kenneth A. Johnson,et al.  A new paradigm for DNA polymerase specificity. , 2006, Biochemistry.

[35]  J. Ninio,et al.  A new approach to DNA polymerase kinetics. , 1979, Journal of molecular biology.

[36]  Ken Garber,et al.  Energy Deregulation: Licensing Tumors to Grow , 2006, Science.

[37]  L. Loeb,et al.  On the fidelity of DNA synthesis:effects of steroids and intercalating agents. , 1980, Chemico-biological interactions.

[38]  S. Benkovic,et al.  Kinetic mechanism whereby DNA polymerase I (Klenow) replicates DNA with high fidelity. , 1988, Biochemistry.

[39]  Oksana Gavrilova,et al.  p53 Regulates Mitochondrial Respiration , 2006, Science.

[40]  Hong Qian,et al.  Phosphorylation energy hypothesis: open chemical systems and their biological functions. , 2007, Annual review of physical chemistry.

[41]  C. Mathews.  Biochemistry of deoxyribonucleic acid-defective amber mutants of bacteriophage T4. 3. Nucleotide pools. , 1972, The Journal of biological chemistry.

[42]  H. Qian.  A simple theory of motor protein kinetics and energetics. , 1997, Biophysical chemistry.

[43]  William Feller,et al.  An Introduction to Probability Theory and Its Applications , 1951 .

[44]  M. Kirschner.  Implications of treadmilling for the stability and polarity of actin and tubulin polymers in vivo , 1980, The Journal of cell biology.

[45]  H. Qian,et al.  Powering a burnt bridges Brownian ratchet: a model for an extracellular motor driven by proteolysis of collagen. , 2006, Physical review. E, Statistical, nonlinear, and soft matter physics.