Investigating Force-Induced Structural Changes in Single Collagen Molecules

Abstract

Collagen is a structural protein found in abundance throughout the body, within the extracellular matrix and connective tissues including skin, bone, and tendons. One of the functions of collagen is to maintain tissue structure in the presence of external forces. However the effect of force on the collagen's structure is not clear:1 in the presence of an external force, collagen has been proposed to have one of two opposing structural changes: either overwinding, reducing its cleavage2 or underwinding, increasing its cleavage.3 Using a high-throughput single-molecule stretching instrument, the centrifuge force microscope (CFM),4 we are developing a technique to investigate force-dependent structural changes of collagen under various loading conditions. Here, we report on our progress tethering, enzymatically cleaving and applying a tunable external force to single molecules of collagen. In addition, we present characterization of the instrument and technique accomplished with single DNA molecules.1.Chang, S.W. and Buehler M. (2014) Molecular biomechanics of collagen molecules. Mat. Today. 17, 70-76.2.Camp, R., Liles, M., Beale, J., Saeidi, N., Flynn, BP., Moore, E., Murthy, S., and Ruberti, JW. (2011) Molecular Mechanochemistry: Low Force Switch Slows Enzymatic Cleavage of Human Type I Collagen Monomer. JACS, 133, 4073-8.3.Adhikari, A., Glassey, E. and Dunn, AR. (2012) Conformation Dynamics Accompanying the Proteolytic Degradation of Trimeric Collagen I by Collagenases. JACS, 134, 13259-65.4.Halvorsen, K. and Wong, W. (2010) Massively Parallel Single-Molecule Manipulation Using Centrifugal Force. Biophys J., 98, L53-L55.