Kessler, Max; Gottschalk, Kay E; Janovjak, Harald LIST Austria ; Mueller, Daniel J; Gaub, Hermann
Despite their crucial importance for cellular function, little is known about the folding mechanisms of membrane proteins. Recently details of the folding energy landscape were elucidated by atomic force microscope (AFM)-based single molecule force spectroscopy. Upon unfolding and extraction of individual membrane proteins energy barriers in structural elements such as loops and helices were mapped and quantified with the precision of a few amino acids. Here we report on the next logical step: controlled refolding of single proteins into the membrane. First individual bacteriorhodopsin monomers were partially unfolded and extracted from the purple membrane by pulling at the C-terminal end with an AFM tip. Then by gradually lowering the tip, the protein was allowed to refold into the membrane while the folding force was recorded. We discovered that upon refolding certain helices are pulled into the membraneagainst a sizable externalforce of several tens of picoNewton. From the mechanical work, which the helix performs on the AFM cantilever, we derive an upper limit for the Gibbs free folding energy. Subsequent unfolding allowed us to analyze the pattern of unfolding barriers and corroborate that the protein had refolded into the native state.
Journal of Molecular Biology
644 - 654
Kessler M, Gottschalk K, Janovjak HL, Mueller D, Gaub H. Bacteriorhodopsin folds into the membrane against an external force. Journal of Molecular Biology. 2006;357(2):644-654. doi:10.1016/j.jmb.2005.12.065
Kessler, M., Gottschalk, K., Janovjak, H. L., Mueller, D., & Gaub, H. (2006). Bacteriorhodopsin folds into the membrane against an external force. Journal of Molecular Biology. Elsevier. https://doi.org/10.1016/j.jmb.2005.12.065
Kessler, Max, Kay Gottschalk, Harald L Janovjak, Daniel Mueller, and Hermann Gaub. “Bacteriorhodopsin Folds into the Membrane against an External Force.” Journal of Molecular Biology. Elsevier, 2006. https://doi.org/10.1016/j.jmb.2005.12.065.
M. Kessler, K. Gottschalk, H. L. Janovjak, D. Mueller, and H. Gaub, “Bacteriorhodopsin folds into the membrane against an external force,” Journal of Molecular Biology, vol. 357, no. 2. Elsevier, pp. 644–654, 2006.
Kessler M, Gottschalk K, Janovjak HL, Mueller D, Gaub H. 2006. Bacteriorhodopsin folds into the membrane against an external force. Journal of Molecular Biology. 357(2), 644–654.
Kessler, Max, et al. “Bacteriorhodopsin Folds into the Membrane against an External Force.” Journal of Molecular Biology, vol. 357, no. 2, Elsevier, 2006, pp. 644–54, doi:10.1016/j.jmb.2005.12.065.