We are using the new and fancy Lumicks Ctrap at Fernando Moreno’ lab (CNB) to track the real-time kinetics of Pfh1 helicase moving along single stranded DNA.
The image above shows the trajectory of a single helicase (blue) tagged with a quantum dot along single-stranded DNA stretched between two optically trapped beads.
This is the result of our collaboration with Roberto Galleto’ lab at Washigton University in St. Louis
As if we were studying the motor of a car, to understand the mechanism of operation of a biological molecular motor we should answer questions like: What moves? Why? What limits performance? What is the amount of force generated? What happen if I pull this?
To answer these questions we are using a single molecule manipulation technique called Optical Tweezers. This novel technique provides previously unobtainable data on fundamental biochemical processes that are essential for all forms of life:
On one hand, the possibility to isolate a single protein and follow its activity in real time allow us to avoid ensemble averaging and capture transient intermediates and heterogeneous behaviour characteristic of the protein operation at the nanos
cale.
On the other hand, we can measure the mechanical forces developed during the protein motor operation and apply controlled forces of a few picoNewtons to it. Access to the mechanical coordinate of the reaction is revealing the mechanisms used by these nanomachines to couple energy to motion.