Manipulation and visualization of single molecules is now possible by means of Magnetic Tweezers. I will introduce this technique in the context of protein machines involved in DNA processing one at a time.
Today, we know that many enzymes and proteins involved in the proper functioning and maintenance of the cell appear in a low-copy number. At this scale level, each molecule counts and proper understanding of a biological process requires monitoring the activity of these individual proteins in action. Moreover, multiple biological processes are time resolved and fundamentally mechanical, following a sequence of events exquisitely coordinated by force, and proving again that life is an out of equilibrium process. Traditional biochemical methods lack this single-molecule approach because they provide information on the ensemble, disregarding individual, out-of-the-mean, behaviors that as mentioned above may be relevant for the proper functioning of the cell. Manipulation and visualization of single molecules is now possible by means of a myriad of biophysical techniques that can capture the presence and activity of single protein machines involved, for instance, in replication, transcription, DNA maintenance and repair, etc. These DNA transactions are essential for the cell, thus it is not surprising that these topics have attracted much attention of researchers over the last years. Here, I will introduce Magnetic Tweezers as a biophysical technique that allow us to investigate protein machines involved in DNA processing one at a time. Magnetic Tweezers is based on an inverted optical microscope and provides a measurement of the extension of a single DNA molecule over time at a given force, while proteins are acting on the DNA. I will show our work of helicases and other proteins involved in DNA repair and chromosome maintenance. Common to all these works is that protein activity is indirectly inferred by changes in the extension of a tethered DNA molecule. This precludes direct identification of the protein acting on the DNA. Prompted by this limitation, we have recently combined Magnetic Tweezers with fluorescence. This state of the art technology opens new possibilities allowing us to correlate biological activity with the identity of DNA-bound protein complexes and to determine their binding position along DNA.
Fecha del seminario: 08/05/2019 12:00
Lugar del seminario: Salón de Actos del Instituto Rocasolano.
Ponente del seminario: Fernando Moreno Herrero
