Direct Visualization of Dynamic Protein-DNA Interactions with a Dedicated Atomic Force Microscope

S.J.T. van Noort, Kees van der Werf, Andre P.M. Eker, Claire Wyman, B.G. de Grooth, N.F. van Hulst, Jan Greve

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Abstract

Photolyase DNA interactions and the annealing of restriction fragment ends are directly visualized with the atomic force microscope (AFM). To be able to interact with proteins, DNA must be loosely bound to the surface. When MgCl2 is used to immobilize DNA to mica, DNA is attached to the surface at distinct sites. The pieces of DNA in between are free to move over the surface and are available for protein interaction. After implementation of a number of instrumental improvements, the molecules can be visualized routinely, under physiological conditions and with molecular resolution. Images are acquired reproducibly without visible damage for at least 30 min, at a scan rate of 2 x 2 mu m(2)/min and a root mean square noise of less than 0.2 nm. Nonspecific photolyase DNA complexes were visualized, showing association, dissociation, and movement of photolyase over the DNA. The latter result suggests a sliding mechanism by which photolyase can scan DNA for damaged sites. The experiments illustrate the potential that AFM presents for modern molecular biology.
Original languageEnglish
Pages (from-to)2840-2849
Number of pages10
JournalBiophysical journal
Volume74
Issue number6
DOIs
Publication statusPublished - 1998

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Deoxyribodipyrimidine Photo-Lyase
DNA
Proteins
Magnesium Chloride
Noise
Molecular Biology
Membrane Proteins

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van Noort, S. J. T., van der Werf, K., Eker, A. P. M., Wyman, C., de Grooth, B. G., van Hulst, N. F., & Greve, J. (1998). Direct Visualization of Dynamic Protein-DNA Interactions with a Dedicated Atomic Force Microscope. Biophysical journal, 74(6), 2840-2849. https://doi.org/10.1016/S0006-3495(98)77991-3
van Noort, S.J.T. ; van der Werf, Kees ; Eker, Andre P.M. ; Wyman, Claire ; de Grooth, B.G. ; van Hulst, N.F. ; Greve, Jan. / Direct Visualization of Dynamic Protein-DNA Interactions with a Dedicated Atomic Force Microscope. In: Biophysical journal. 1998 ; Vol. 74, No. 6. pp. 2840-2849.
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abstract = "Photolyase DNA interactions and the annealing of restriction fragment ends are directly visualized with the atomic force microscope (AFM). To be able to interact with proteins, DNA must be loosely bound to the surface. When MgCl2 is used to immobilize DNA to mica, DNA is attached to the surface at distinct sites. The pieces of DNA in between are free to move over the surface and are available for protein interaction. After implementation of a number of instrumental improvements, the molecules can be visualized routinely, under physiological conditions and with molecular resolution. Images are acquired reproducibly without visible damage for at least 30 min, at a scan rate of 2 x 2 mu m(2)/min and a root mean square noise of less than 0.2 nm. Nonspecific photolyase DNA complexes were visualized, showing association, dissociation, and movement of photolyase over the DNA. The latter result suggests a sliding mechanism by which photolyase can scan DNA for damaged sites. The experiments illustrate the potential that AFM presents for modern molecular biology.",
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van Noort, SJT, van der Werf, K, Eker, APM, Wyman, C, de Grooth, BG, van Hulst, NF & Greve, J 1998, 'Direct Visualization of Dynamic Protein-DNA Interactions with a Dedicated Atomic Force Microscope' Biophysical journal, vol. 74, no. 6, pp. 2840-2849. https://doi.org/10.1016/S0006-3495(98)77991-3

Direct Visualization of Dynamic Protein-DNA Interactions with a Dedicated Atomic Force Microscope. / van Noort, S.J.T.; van der Werf, Kees; Eker, Andre P.M.; Wyman, Claire; de Grooth, B.G.; van Hulst, N.F.; Greve, Jan.

In: Biophysical journal, Vol. 74, No. 6, 1998, p. 2840-2849.

Research output: Contribution to journalArticleAcademicpeer-review

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AU - van der Werf, Kees

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AU - de Grooth, B.G.

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AU - Greve, Jan

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AB - Photolyase DNA interactions and the annealing of restriction fragment ends are directly visualized with the atomic force microscope (AFM). To be able to interact with proteins, DNA must be loosely bound to the surface. When MgCl2 is used to immobilize DNA to mica, DNA is attached to the surface at distinct sites. The pieces of DNA in between are free to move over the surface and are available for protein interaction. After implementation of a number of instrumental improvements, the molecules can be visualized routinely, under physiological conditions and with molecular resolution. Images are acquired reproducibly without visible damage for at least 30 min, at a scan rate of 2 x 2 mu m(2)/min and a root mean square noise of less than 0.2 nm. Nonspecific photolyase DNA complexes were visualized, showing association, dissociation, and movement of photolyase over the DNA. The latter result suggests a sliding mechanism by which photolyase can scan DNA for damaged sites. The experiments illustrate the potential that AFM presents for modern molecular biology.

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van Noort SJT, van der Werf K, Eker APM, Wyman C, de Grooth BG, van Hulst NF et al. Direct Visualization of Dynamic Protein-DNA Interactions with a Dedicated Atomic Force Microscope. Biophysical journal. 1998;74(6):2840-2849. https://doi.org/10.1016/S0006-3495(98)77991-3