Combining optical tweezers and scanning probe microscopy to study DNA-protein interactions

J.H.G. Huisstede, Vinod Subramaniam, Martin L. Bennink

Research output: Contribution to journalArticleAcademicpeer-review

16 Citations (Scopus)

Abstract

We present the first results obtained with a new instrument designed and built to study DNA-protein interactions at the single molecule level. This microscope combines optical tweezers with scanning probe microscopy and allows us to locate DNA-binding proteins on a single suspended DNA molecule. A single DNA molecule is stretched taut using the optical tweezers, while a probe is scanned along the molecule. Interaction forces between the probe and the sample are measured with the optical tweezers. The instrument thus enables us to correlate mechanical and functional properties of bound proteins with the tension within the DNA molecule. The typical friction force between a micropipette used as probe and a naked DNA molecule was found to be <1 pN. A 16 m DNA molecule with 10-15 digoxygenin (DIG) molecules located over a 90 nm range in the middle of the DNA was used as a model system. By scanning with an antidigoxygenin (-DIG) antibody-coated pipette we were able to localize these sites by exploiting the high binding affinity between this antibody-antigen pair. The estimated experimental resolution assuming an infinitesimally thin and rigid probe and a single -DIG/DIG bond was 15 nm. Microsc. Res. Tech., 2006.
Original languageUndefined
Pages (from-to)26-33
Number of pages8
JournalMicroscopy research and technique
Volume70
Issue number1
DOIs
Publication statusPublished - 2007

Keywords

  • single molecule force spectroscopy
  • digoxygenin
  • Optical tweezers
  • DNA-protein interactions
  • Scanning probe microscopy
  • IR-72338
  • METIS-241379

Cite this

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title = "Combining optical tweezers and scanning probe microscopy to study DNA-protein interactions",
abstract = "We present the first results obtained with a new instrument designed and built to study DNA-protein interactions at the single molecule level. This microscope combines optical tweezers with scanning probe microscopy and allows us to locate DNA-binding proteins on a single suspended DNA molecule. A single DNA molecule is stretched taut using the optical tweezers, while a probe is scanned along the molecule. Interaction forces between the probe and the sample are measured with the optical tweezers. The instrument thus enables us to correlate mechanical and functional properties of bound proteins with the tension within the DNA molecule. The typical friction force between a micropipette used as probe and a naked DNA molecule was found to be <1 pN. A 16 m DNA molecule with 10-15 digoxygenin (DIG) molecules located over a 90 nm range in the middle of the DNA was used as a model system. By scanning with an antidigoxygenin (-DIG) antibody-coated pipette we were able to localize these sites by exploiting the high binding affinity between this antibody-antigen pair. The estimated experimental resolution assuming an infinitesimally thin and rigid probe and a single -DIG/DIG bond was 15 nm. Microsc. Res. Tech., 2006.",
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Combining optical tweezers and scanning probe microscopy to study DNA-protein interactions. / Huisstede, J.H.G.; Subramaniam, Vinod; Bennink, Martin L.

In: Microscopy research and technique, Vol. 70, No. 1, 2007, p. 26-33.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Combining optical tweezers and scanning probe microscopy to study DNA-protein interactions

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AU - Subramaniam, Vinod

AU - Bennink, Martin L.

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Y1 - 2007

N2 - We present the first results obtained with a new instrument designed and built to study DNA-protein interactions at the single molecule level. This microscope combines optical tweezers with scanning probe microscopy and allows us to locate DNA-binding proteins on a single suspended DNA molecule. A single DNA molecule is stretched taut using the optical tweezers, while a probe is scanned along the molecule. Interaction forces between the probe and the sample are measured with the optical tweezers. The instrument thus enables us to correlate mechanical and functional properties of bound proteins with the tension within the DNA molecule. The typical friction force between a micropipette used as probe and a naked DNA molecule was found to be <1 pN. A 16 m DNA molecule with 10-15 digoxygenin (DIG) molecules located over a 90 nm range in the middle of the DNA was used as a model system. By scanning with an antidigoxygenin (-DIG) antibody-coated pipette we were able to localize these sites by exploiting the high binding affinity between this antibody-antigen pair. The estimated experimental resolution assuming an infinitesimally thin and rigid probe and a single -DIG/DIG bond was 15 nm. Microsc. Res. Tech., 2006.

AB - We present the first results obtained with a new instrument designed and built to study DNA-protein interactions at the single molecule level. This microscope combines optical tweezers with scanning probe microscopy and allows us to locate DNA-binding proteins on a single suspended DNA molecule. A single DNA molecule is stretched taut using the optical tweezers, while a probe is scanned along the molecule. Interaction forces between the probe and the sample are measured with the optical tweezers. The instrument thus enables us to correlate mechanical and functional properties of bound proteins with the tension within the DNA molecule. The typical friction force between a micropipette used as probe and a naked DNA molecule was found to be <1 pN. A 16 m DNA molecule with 10-15 digoxygenin (DIG) molecules located over a 90 nm range in the middle of the DNA was used as a model system. By scanning with an antidigoxygenin (-DIG) antibody-coated pipette we were able to localize these sites by exploiting the high binding affinity between this antibody-antigen pair. The estimated experimental resolution assuming an infinitesimally thin and rigid probe and a single -DIG/DIG bond was 15 nm. Microsc. Res. Tech., 2006.

KW - single molecule force spectroscopy

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KW - DNA-protein interactions

KW - Scanning probe microscopy

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