Comparison between 102k and 20k Poly(ethylene oxide) Depletants in Osmotic Pressure Measurements of Interfilament Forces in Cytoskeletal Systems,ACS Macro Letters

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Comparison between 102k and 20k Poly(ethylene oxide) Depletants in Osmotic Pressure Measurements of Interfilament Forces in Cytoskeletal Systems
ACS Macro Letters ( IF 5.1 ) Pub Date : 2018-02-01 00:00:00 , DOI: 10.1021/acsmacrolett.7b00937
Peter J Chung , Chaeyeon Song , Joanna Deek , Herbert P Miller , Youli Li , Myung Chul Choi ₁ , Leslie Wilson , Stuart C Feinstein , Cyrus R Safinya

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The proliferation of successful, cell-free reconstitutions of cytoskeletal networks have prompted measurements of forces between network elements via induced osmotic pressure by the addition of depletants. Indeed, it was through osmotic pressurization that Tau, an intrinsically disordered protein found in neuronal axons, was recently discovered to mediate two distinct microtubule (MT) bundle states, one widely spaced and a second tightly packed, separated by an energy barrier due to polyelectrolyte repulsions between opposing Tau projection domains on neighboring MT surfaces. Here, we compare interfilament force measurements in Tau coated MT bundles using PEO20k (poly(ethylene oxide), M_w = 20000), a commonly used inert depletant, and recently published measurements with PEO102k. While force measurements with either depletant reveals the transition between the two bundled states, measurements with PEO20k cannot recapitulate the correct critical pressure (P_c) at which widely spaced MT bundles transition to tightly packed MT bundles due to depletant penetration into widely spaced bundles below P_c. Surprisingly, upon transitioning to the tightly packed bundle state data from both depletants are in quantitative agreement indicative of expulsion of the smaller PEO20k depletant, but only at distances comparable or less than the PEO20k radius of gyration, significantly smaller than the effective diameter of PEO20k. While PEO102k (with size larger than the wall-to-wall distance between MTs in bundles) can more accurately capture the force response behavior at low to intermediate pressures (<10⁴ Pa), measurements with PEO20k, beyond the overlap regime with PEO102k, extend the achievable osmotic pressure range into the higher-pressure regime (∼5 × 10⁴ Pa). The data underscores the importance of the use of polymeric depletants of different sizes to elucidate force response behavior of cytoskeletal filamentous networks over a more complete extended pressure range.

中文翻译：

102k 和 20k 聚（环氧乙烷）消耗剂在细胞骨架系统中丝间力渗透压测量中的比较

细胞骨架网络的成功、无细胞重组的扩散促进了通过添加消耗剂诱导的渗透压来测量网络元素之间的力。事实上，最近发现 Tau 是一种在神经元轴突中发现的本质上无序的蛋白质，它通过渗透压来介导两种不同的微管 (MT) 束状态，一种是间隔广泛的，另一种是紧密排列的，由聚电解质引起的能量屏障隔开。相邻 MT 表面上相对的 Tau 投影域之间的排斥。在这里，我们比较了使用 PEO20k（聚（环氧乙烷），M _w= 20000)，一种常用的惰性消耗剂，最近发表了使用 PEO102k 进行的测量。虽然使用任一消耗剂的力测量揭示了两种捆绑状态之间的转变，但使用 PEO20k 的测量无法概括正确的临界压力 ( _Pc) ，在该压力下，由于消耗剂渗透到低于P的宽间距束中，宽间距 MT 束转变为紧密包装的 MT 束_C. 令人惊讶的是，在转换到紧密堆积的束状态时，两种消耗剂的数据在定量上一致，表明较小的 PEO20k 消耗剂被排出，但仅在与 PEO20k 回转半径相当或小于的距离处，明显小于 PEO20k 的有效直径。虽然 PEO102k（尺寸大于束中 MT 之间的壁到壁距离）可以更准确地捕捉在低至中压（<10 ⁴ Pa）下的力响应行为，但使用 PEO20k 进行的测量，超出了使用 PEO102k 的重叠范围，将可达到的渗透压范围扩展到更高的压力范围（~5 × 10 ⁴帕）。这些数据强调了使用不同尺寸的聚合物消耗剂在更完整的扩展压力范围内阐明细胞骨架丝状网络的力响应行为的重要性。

更新日期：2018-02-01

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