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    Scientists Use Laser ‘Tweezers’ to Grab, Study Protein Droplets

    Article obtained from Photonics RSS Feed.

    University at Buffalo physicists are using lasers to study proteins that cluster together to form spherical droplets inside human cells, shedding light on the conditions that drive such droplets to switch from a fluid, liquidy state to a harder, gel-like state.

    The study finds that certain protein droplets harden, becoming gelatinous in crowded environments, such as test tubes where many other molecules are present, thereby mimicking the congested conditions inside living cells.

    Lead investigator Priya R. Banerjee, assistant professor of physics in the College of Arts and Sciences, said that very little is known about the droplet-forming proteins’ basic properties, leading physicists to quantify the dynamics of these droplets and learn what factors influence them. This is important as the dynamics of protein droplets are a key to their cellular function and dysfunction.

    “Prior research has focused on the structure of the proteins themselves, but our work shows that environmental factors are equally important,” Banerjee said. “We see that external conditions can alter the internal state of the droplets, which may affect their function in human cells.”

    The research, she added, matters because condensating proteins may be involved in health and disease. Recent studies point to potential roles for these droplets in such diverse functions as gene expression, stress response, and immune system function.

    The research investigates a droplet-forming protein called fused in sarcoma (FUS). Liquid FUS droplets are found in normal brain cells, but in some patients with the neurodegenerative disease amyotrophic lateral sclerosis (ALS), the protein forms aggregates of solid material, Banerjee said.
                    Micro droplets. Courtesy of Priya Banerjee, Lab at UB.
     The research employed two innovative laser techniques to show how environmental conditions can affect droplets made from FUS or other related proteins.

    In one set of experiments, scientists used highly focused laser beams called optical tweezers to trap and push together two protein droplets floating in a liquid buffer solution. The droplets merged easily to form a single, larger droplet when the buffer was thinly populated with other inert crowder molecules such as polyethylene glycol (PEG). But when the concentration of PEG or other chemicals in the buffer increased, the protein droplets became more gelatinous and would not fully combine.

    In a second set of tests, the team employed lasers in a different way using “laser poking” to study how FUS and related protein droplets react to crowded environments.

    In these experiments, Banerjee and colleagues attached fluorescent tags to numerous protein molecules in a single droplet, causing the proteins to glow. The researchers then “poked” the middle of the droplet with a high-intensity laser, a procedure that caused any fluorescent molecules hit by the laser to go permanently dark.

    The next step in the process was for scientists to measure how long it took for new glowing proteins to move into the darkened area. This happened quickly in protein droplets floating in sparsely populated buffer solutions. But the recovery time was dramatically slower for droplets suspended in buffer solutions thick with PEG or other compounds, an indication that protein droplets become gelatinous in crowded environments. The findings applied to both FUS and other related protein droplets with diverse primary structures.

    “Our experiments were done in test tubes, but our results suggest that inside living cells, the crowding status could affect the dynamics of protein droplets,” Banerjee said.

    One important question that remains is whether and how the fluidity of FUS droplets affects the protein’s ability to form into solid clumps, as seen in some ALS patients. Banerjee hopes to address this problem through future research.

    The study in biomolecules was supported by the College of Arts and Sciences, with assistance from the University at Buffalo North Campus Confocal Imaging Facility, which is supported by the National Science Foundation. The research was conducted by Banerjee’s team at UB, with technical assistance from a colleague affiliated with Baylor College of Medicine.
     

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    Mar, 11 2019 |

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