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(530190) Methods in Single Molecule Biophysics, (3op) , Fall 2011
The central dogma states that information in nucleic acid (DNA in cells) can be perpetuated (replication) or transferred (transcription), but the transfer of information form into protein (translation) is irreversible.
Why single molecules?
• no need for sample synchronization
• detection of intermediates
• single molecule measurements avoid ensemble averaging
• full statistical description can be achieved
• When does averaging work? • Do we need to observe a single molecule ?
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Why single molecule biology?
Does nature look like this?
... Or this?
Why Single Molecule Biophysics ?
• Ensemble measurements give: Averages over large populations (+errors for these averages)
• Single Molecule measurements give:Distributions ! • Rare events!
Adelman et. al., PNAS, 2002, Vol 99 no 21
Single Molecule Biology • DNA
– What kind of a polymer is DNA ? – How do DNA-enzymes work ? – DNA-compaction
• The folding problem – Proteins – RNA
• Molecular motors/machines – How do they work? – How is chemical energy converted to mechanical
• hydrolysis energy ~ 20 kBT = 80 pN nm Hoenger et al., J Mol Biol, 297, 1087-103
Examples of Molecular Motors
Myosin is present in human skeletal muscle Myosin steps along an Actin track Kinesin transports various cargos (vesicles) inside the cell Kinesin steps along a Mircotubule track
Videos of optical tweezers
• Stretching DNA in optical tweezers: http://www.youtube.com/watch?v=HsuwMnMIEyc
• Lambda exonuclease, a simple molecular motor: http://www.youtube.com/watch?v=ddPdxk6-1k0