A nanoplasmonic molecular ruler for measuring nuclease activity and DNA footprinting

Abstract

Interactions between nucleic acids and proteins are essential to genetic information processing. The detection of size changes in nucleic acids is the key to mapping such interactions, and usually requires substrates with fluorescent, electrochemical or radioactive labels1,2,3. Recently, methods have been developed to tether DNA to highly water-soluble Au nanoparticles4,5,6,7,8, and nanoparticle pairs linked by DNA have been used to measure nanoscale distances9. Here we demonstrate a molecular ruler in which double-stranded DNA is attached to a Au nanoparticle. The change in plasmon resonance wavelength of individual Au–DNA conjugates depends on the length of the DNA and can be measured with subnanometre axial resolution. An average wavelength shift of approximately 1.24 nm is observed per DNA base pair. This system allows for a label-free, quantitative, real-time measurement of nuclease activity and also serves as a new DNA footprinting platform, which can accurately detect and map the specific binding of a protein to DNA.

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Figure 1: Design of the Au–DNA nanoplasmonic molecular ruler.
Figure 2: DNA conjugation red-shifted plasmon resonance peak of the molecular ruler.
Figure 3: Endonuclease activity measurement with ruler.
Figure 4: Calibration curve generated by nanoplasmon resonance sensing of multiple enzymes.
Figure 5: DNA footprinting of Bal31 exonuclease stalled by the EcoRI(Q111) proteins.

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Acknowledgements

This work was supported by DARPA, DOD BC045345, NIH R21CA95393, UCSF Prostate Cancer SPORE award (NIH P50 CA89520), and the UCSF Prostate Cancer Center Developmental Research Program, Intel, the Korea Ministry of Science and Technology “21st Century Frontier R&D Program” grant 05K1501-02810. This work was performed under the auspices of the U.S. Dept. of Energy, at the University of California/Lawrence Berkeley National Laboratory under contract no. DE-AC03-76SF00098 and at the University of California/Lawrence Livermore National Laboratory under contract no. W-7405-Eng-48.

Author information

F.F.C. conceived and designed the experiments, G.L.L., Y.Y., S.K. and B.M. performed the experiments, G.L.L. and D.G. analysed the data, and S.D.J. and D.G.B. synthesized the EcoRI(Q111). Correspondence and requests for material should be addressed to F.F.C. and L.P.L.

Correspondence to Luke P. Lee or Fanqing Frank Chen.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary materials and methods, figures S1-S4, and table S1 (PDF 254 kb)

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Liu, G., Yin, Y., Kunchakarra, S. et al. A nanoplasmonic molecular ruler for measuring nuclease activity and DNA footprinting. Nature Nanotech 1, 47–52 (2006). https://doi.org/10.1038/nnano.2006.51

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