Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 106, Issue 4, Pages 979-984Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.0812356106
Keywords
DNA machine; synthetic biology
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Funding
- National Science Foundation (NSF) [DMR-0213618]
- National Institutes of Health [GM069630]
- Materials Research Science and Engineering Centers
- Div Of Molecular and Cellular Bioscience
- Direct For Biological Sciences [0918782] Funding Source: National Science Foundation
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Synthetic lipid-oligonucleotide conjugates inserted into lipid vesicles mediate fusion when one population of vesicles displays the 5'-coupled conjugate and the other the 3'-coupled conjugate, so that anti-parallel hybridization allows the membrane surfaces to come into close proximity. Improved assays show that lipid mixing proceeds more quickly and to a much greater extent than content mixing, suggesting the latter is rate limiting. To test the effect of membrane-membrane spacing on fusion, a series of conjugates was constructed by adding 2-24 noncomplementary bases at the membrane-proximal ends of two complementary sequences. Increasing linker lengths generally resulted in progressively reduced rates and extents of lipid and content mixing, in contrast to higher vesicle docking rates. The relatively flexible, single-stranded DNA linker facilitates docking but allows greater spacing between the vesicles after docking, thus making the transition into fusion less probable, but not preventing it altogether. These experiments demonstrate the utility of DNA as a model system for fusion proteins, where sequence can easily be modified to systematically probe the effect of distance between bilayers in the fusion reaction.
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