Generating enzyme-synthesized hyaluronan brushes: Characterization, length regulation and applications

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Wei, Wenbin
Curtis, Jennifer E.
Gumbart, James C.
Kim, Harold D.
Brettmann, Blair K.
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Polymer brushes are tethered polymers at an interface with a high grafting density. Polymer brushes have rich physics phenomena and wide applications. We developed a distinct class of polymer brush which increased the typical length scale of polymer brush thickness by an order of magnitude, from less than a few hundred nanometers100 nm to ten microns. Hyaluronan polymers were polymerized and extruded by the enzyme hyaluronan synthase, which was densely coated on a surface. The resultant hyaluronan brush can be visualized using optical microscopy which makes them a unique tool to study the physics of polymer brushes in ways previously never accessible. We developed a toolkit to study the interactions between the hyaluronan brush and proteins, nanoparticles, bacteria and mammalian cells. We found the hyaluronan polymers, not the enzymes, serve as the memory of the length regulation mechanism which may result in longer polymers detaching at a faster rate than shorter polymers. In addition to the rich physics in the system, hyaluronan brushes are a promising biomaterial. Hyaluronan is an important and abundant biological polymer as an adult carries 12-15 g hyaluronan, 1/3 of which is degraded and synthesized every day. In testing its potential in the application of implants, we found the brush allowed normal attachment and proliferation of mammalian fibroblast cells but that blocks a strain of bacteria which is the most frequent pathogen associated with implants for at least two weeks. Our initial and most ambitious goal was to test the hypothesis that polymer physics provides a length-dependent tension on the enzyme-bound polymers. This would be a natural explanation of the length-dependent detachment rates observed. Although this ultimate goal is still under pursuit, we have established an excellent platform to address the question and in the process, opened unexpected but valuable avenues of research in polymer physics, cell biophysics, and antimicrobial biomaterials.
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