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Single-Molecule Kinetics of λ Exonuclease Reveal Base Dependence and Dynamic Disorder
Open accessAbstract
We used a multiplexed approach based on flow-stretched DNA to monitor the enzymatic digestion of λ-phage DNA by individual bacteriophage λ exonuclease molecules. Statistical analyses of multiple single-molecule trajectories observed simultaneously reveal that the catalytic rate is dependent on the local base content of the substrate DNA. By relating single-molecule kinetics to the free energies of hydrogen bonding and base stacking, we establish that the melting of a base from the DNA is the rate-limiting step in the catalytic cycle. The catalytic rate also exhibits large fluctuations independent of the sequence, which we attribute to conformational changes of the enzyme-DNA complex.
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Cited by
Direct observation of base-pair stepping by RNA polymerase
Abbondanzieri, E. A., Greenleaf, W. J., Shaevitz, J. W., Landick, R., & Block, S. M. (2005) · Nature · doi:10.1038/nature04268 · Cited by 855
Statistical Kinetics of Macromolecular Dynamics
Shaevitz, J. W., Block, S. M., & Schnitzer, M. J. (2005) · Biophysical Journal · doi:10.1529/biophysj.105.064295 · Cited by 86
Direct observation of processive exoribonuclease motion using optical tweezers
Fazal, F. M., Koslover, D. J., Luisi, B. F., & Block, S. M. (2015) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1514028112 · Cited by 29
Vannoy, K. J., Ryabykh, A., Chapoval, A. I., & Dick, J. E. (2021) · The Analyst · doi:10.1039/d1an00230a · Cited by 26
Metadata
- Authors
- van Oijen, A. M., Blainey, P. C., Crampton, D. J., Richardson, C. C., Ellenberger, T., & Xie, X. S.
- Venue
- Science
- Published
- 2003-08-29
- Volume
- 301
- Issue
- 5637
- Pages
- 1235-1238
- ISSN
- 0036-8075 (ISSN-L)
- 1095-9203
- OpenAlex
- W2131448623
- Cited by
- 372
- References
- 0
- FWCI
- 7.08
- Type
- journal article
- Language
- en
- Open access
- green
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- van Oijen, A. M., Blainey, P. C., Crampton, D. J., Richardson, C. C., Ellenberger, T., & Xie, X. S. (2003). Single-Molecule Kinetics of λ Exonuclease Reveal Base Dependence and Dynamic Disorder. Science. https://doi.org/10.1126/science.1084387
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