Scanning probe optical tweezers

Dr. Jurgen Huisstede (BEP group)

Background

The functions of all biological processes within the cell are regulated by specific interactions of proteins with nucleic acids or other proteins controlling processes such as transcription, translation, repair and replication that are essential for all life. To understand these processes several single molecule techniques are developed to visualize or manipulate these processes.

Objective

A new microscope, the scanning probe optical tweezers, has been developed and built to study DNA-protein interactions at the single molecule level. Combining a reflection-based optical tweezers with scanning probe microscopy we want to correlate positional information of proteins bound to a single DNA molecule with biomechanical transitions that occur due to specific DNA-protein interactions.

A single DNA molecule is stretched and free from any surface, except at both its ends where it is coupled to a bead or surface. In a next step the stretched DNA molecule with proteins will be approached by a probe. This probe, a multi-walled carbon nanotube with a diameter of 40-90 nm, will scan back and forth to “feel” the individual proteins as it touches them. In contrast to conventional SPM techniques, three dimensional diffusive processes are not obstructed by a supporting surface except by the proximity of the probe, through which the system better resembles conditions as in vivo. Yet a small stretching force is required for localization. A drawback for the experiments is the many steps that are required to attach a single DNA molecule between two beads and to start scanning with the probe, making the experiments time-consuming. Acquiring statistics is therefore a difficult and lengthy process. Nevertheless the SPOT microscope provides valuable information with several potential applications.

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