Nanometer-scale protein molecular machines are at the core of all living systems. For example, helicase motor proteins consume ATP fuel molecules and to walk down DNA tightropes unwinding the double helix. This activity is essential for e.g., allowing access to the information stored in DNA or repairing damaged DNA. How these machines function remains mysterious.
The Comstock lab investigates such processes using advanced, precision single-molecule measurement and manipulation techniques. Using optical traps we can grab and stretch a single DNA strand and observe the activity of individual protein molecular machines in real-time (see cartoon on right).
Amongst other projects, we are presently collaborating with the Jankowsky lab at Case Western Reserve University to investigate molecular machine processing of RNA.
We have recently published results from this project where we have directly shown that the nuclear RNA helicase Mtr4p (both alone and within the TRAMP RNA surveilance complex) is a slow translocase that stalls without an upstream duplex to unwind:E. M. Patrick, Sukanya Srinivasan, Eckhard Jankowsky, and M. J. Comstock. “The RNA Helicase Mtr4p is a Duplex Sensing Translocase.” Nature Chemical Biology, 13, 99 (2017).
For more information on our technique see:
M.J. Comstock, K.D. Whitley, H. Jia, J. Sokoloski, T.M. Lohman, Taekjip Ha, and Y.R. Chemla. “Direct observation of structure-function relationship in a nucleic acid-processing enzyme,” Science, 348, 352 (2015).
Protocols and methods book chapters:
K. D. Whitley, M. J. Comstock, Y. R. Chemla. "High-resolution optical tweezers combined with single-molecule confocal microscopy," in “Methods in Enzymology”: “Single-Molecule Enzymology: Nanomechanical Manipulation and Hybrid Methods”, Spies, M. & Chemla, Y. R., (editors), Elsevier. 582 (2017).
K. D. Whitley, M. J. Comstock, Y. R. Chemla. "High-Resolution “Fleezers”: Dual-trap Optical Tweezers Combined with Single-Molecule Fluorescence Detection," in: “Optical Tweezers: Methods and Protocols”, Generich, A. (editor), Springer, (2016, in press).
The optical traps can be configured to tug on the ends of individual proteins to induce the unraveling and refolding of their structure. We are presently collaborating with the Lapidus lab to investigate protein folding processes using our novel high-resolution trapping and fluorescence capabilities.
Trapping and probing individual cells
We also use the optical traps to grab onto individual live cells and precisely manipulate and measure their properties. We are presently collaborating with the Reguera lab at MSU to measure the biological fuel cell capabilities of individual cells.