Berlin 2005 – scientific programme

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AKB: Biologische Physik

AKB 200: Poster Session II

AKB 200.1: Poster

Tuesday, March 8, 2005, 17:00–19:00, Poster TU C

Probing of the proteasom-protein interaction with force-spectroscopy — •Mirjam Beuttler, Jens Schiener, and Reinhard Guckenberger — Max-Planck-Institut für Biochemie, 82152 Martinsried

Atomic force microscopy (AFM) is an established method to investigate biological samples in their physiological environment. In our group we are investigating the 20S proteasome from Thermoplasma acidophilum. Besides imaging we will focus on force-measuring.

The proteasome is a barrel-shaped enzyme of 15 nm height and 11 nm width with a small opening at both ends. Through these two entrances unfolded proteins can access the inner part of the proteasome with the catalytic centers in order to be degraded there. Our goal is to characterize the translocation mechanism, as the forces involved are currently unknown. First step is to immobilize the proteasomes in an upright position which is achieved in our case directly on mica. Imaging the samples ensures the right orientation and the density of the surface covering. Second step will be the investigation of the forces involved in the translocation mechanism. Therefore suitable proteins which are known to be degraded by the proteasome will be bound to the AFM-tip. The forces exerted on the proteins by the proteasomes are transmitted to the tip. While retracting the lever with the tip from the surface the deflection of the lever changes due to the forces. Similarly, when the lever is kept stationary it will be bent towards the sample when the protein is sucked into the proteasome. Such force-distance-curves are offering a promising route to a better understanding of the translocation mechanism.