The elegant architecture of the channel of bacteriophage phi29 DNA packaging motor has inspired the development of biomimetics for biophysical and nanobiomedical applications. The reengineered channel inserted into a lipid membrane exhibits robust electrophysiological properties ideal for precise sensing and fingerprinting of dsDNA at the single-molecule level. Herein, we used single channel conduction assays to quantitatively evaluate the translocation dynamics of dsDNA as a function of the length and conformation of dsDNA. We extracted the speed of dsDNA translocation from the dwell time distribution and estimated the various forces involved in the translocation process. A ~35-fold slower speed of translocation per base-pair was observed for long dsDNA, a significant contrast to the speed of dsDNA crossing synthetic pores. It was found that the channel could translocate both dsDNA with ~32% of channel current blockage and with ~64% for tetra-stranded DNA (two parallel dsDNA). The calculation of both cross-sectional areas of the dsDNA and tetra-stranded DNA suggested that the blockage was purely proportional to the physical space of the channel lumen and the size of the DNA substrate. Folded dsDNA configuration was clearly reflected in their characteristic current signatures. The finding of translocation of tetra-stranded DNA with 64% blockage is in consent with the recently elucidated mechanism of viral DNA packaging via a revolution mode that requires a channel larger than the dsDNA diameter of 2nm to provide room for viral DNA revolving without rotation. The understanding of the dynamics of dsDNA translocation in the phi29 system will enable us to design more sophisticated single pore DNA translocation devices for future applications in nanotechnology and personal medicine.
|Number of pages||9|
|State||Published - Jun 1 2015|
Bibliographical noteFunding Information:
We thank Dr. Hui Zhang and Dr. Mario Vieweger for help with the data analysis. The work was supported by NIH grant EB012135 (P.G.) and, National Cancer Institute Cancer Center Support Grant P30CA177558 . Funding to Peixuan Guo's Endowed Chair in Nanobiotechnology position is from the William Fairish Endowment Fund . Peixuan Guo is a co-founder of Kylin Therapeutics, Inc., and Biomotor and RNA Nanotechnology Development Corp. Ltd.
© 2015 Elsevier Ltd.
- DNA conformation
- Phi29 connector
- Single channel conductance
ASJC Scopus subject areas
- Ceramics and Composites
- Mechanics of Materials