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Proceedings Paper

Protein folding and unfolding by force microscopy: a novel inverse methodology
Author(s): Gregory Shaun Watson; Jolanta Anna Watson; Christopher L. Brown; Sverre Myhra
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Paper Abstract

It has been shown recently that it is possible to reverse-engineer the folding of complex proteins by an AFM-based force versus distance methodology. In essence, the protein is attached to a functionalised surface, then linked to a functionalized tip of a force-sensing/imposing lever, and finally stretched by withdrawal of the probe. The experiments are technically demanding, and subject to a number of artefacts. Moreover, the unfolding process cannot readily be reversed. A 'reverse’ methodology is possible in principle and is likely to provide greater insight into the folding/unfolding sequence. The protein in question will now be attached to a functionalized superparamagnetic bead in solution. A functionalized probe is then introduced into solution whereupon the protein (and its bead) attaches itself to the tip of the probe, while being tracked optically. The system is then submerged in an inhomogeneous magnetic dipole field, giving rise to a magnetic dipole-dipole interaction with the bead. The resultant force will stretch the protein, while being monitored at a force resolution of ca. 1 pN by the deflection of the lever. The significant aspect of the scheme is that the probe acts as a position-sensitive element as well as being a force-sensing device. The field gradient and its rate of change within the interaction volume can readily be controlled to 1 part in 106, and can be reversed. Thus the unfolding/folding sequence can be retraced for the same molecule. Moreover, information about kinetics will be accessible from varying the rate of change in field strength, and the effects of ambient fluid conditions can be investigated in real time with a flow-through arrangement. The proposed methodology is likely to be more userfriendly as a result of dispensing with the functionalized surface, and by being able to determine optically that a single functionalized bead has arrived at the probe tip. Elements of the experimental arrangement are currently at the design stage.

Paper Details

Date Published: 23 February 2005
PDF: 8 pages
Proc. SPIE 5650, Micro- and Nanotechnology: Materials, Processes, Packaging, and Systems II, (23 February 2005); doi: 10.1117/12.582646
Show Author Affiliations
Gregory Shaun Watson, Griffith Univ. (Australia)
Jolanta Anna Watson, Griffith Univ. (Australia)
Christopher L. Brown, Griffith Univ. (Australia)
Sverre Myhra, Griffith Univ. (Australia)
Univ. of Oxford (United Kingdom)

Published in SPIE Proceedings Vol. 5650:
Micro- and Nanotechnology: Materials, Processes, Packaging, and Systems II
Jung-Chih Chiao; David N. Jamieson; Lorenzo Faraone; Andrew S. Dzurak, Editor(s)

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