A new tool for measuring surface-biomolecule interactions

项目摘要。为什么生物分子在附着在某些表面（例如，细胞 膜）并且当附着于其它膜时几乎总是展开和折叠（例如，许多人工 表面）？我们如何才能重现前一种效果，制造出人造表面， 生物分子同样保持其功能？到目前为止，我们解决这些重要问题的能力一直是 由于严重缺乏定量的实验方法来测量热力学， 生物分子-表面相互作用。也就是说，尽管大量定性文献描述了如何 吸附改变了生物分子结构，大量的实验研究寻找吸附- 抗性表面，定量，实验测试的见解如何以及为什么会发生这种情况已经证明 难以捉摸。因此，出于动机，我们建议在这里开发和验证一种新的实验 （电化学）方法来测量生物分子的折叠自由能的位点特异性连接到 清晰的宏观表面。然后与本体溶液中的折叠自由能进行比较， 生物聚合物-表面相互作用的热力学机制。迄今为止， 采用这种方法来表征表面限制的DNA茎环的容易建模的折叠， 这些研究第一次通过实验确定了一个人在多大程度上和在多大程度上 特定生物分子与一系列明确的宏观表面相互作用。在此，我们提出了一个为期两年的 一项研究计划，旨在使这种定量实验工具适用于蛋白质表面的研究。 交互.

Project summary. Why do biomolecules retain activity when attached to some surfaces (e.g., the cell membrane) and yet almost invariably unfold and inactivate when attached to others (e.g., many artificial surfaces)? And how can we recreate the former effect to produce artificial surfaces on which attached biomolecules similarly retain their function? To date our ability to address these important questions has been hampered by an acute lack of quantitative experimental methods for measuring the thermodynamics of biomolecule-surface interactions. That is, despite a large body of qualitative literature describing how adsorption alters biomolecular structure, and a large number of empirical studies searching for adsorption- resistant surfaces, quantitative, experimentally testable insights into how and why this occurs have proven elusive. Thus motivated, we propose here the development and validation of a novel experimental (electrochemical) approach to measuring the folding free energy of biomolecules site-specifically attached to well-defined macroscopic surfaces. Comparison with the folding free energy in bulk solution then informs on the thermodynamics -and thus mechanisms- underlying biopolymer-surface interactions. To date we have employed this approach to characterize the easily modeled folding of a surface-confined DNA stem-loop in studies that have, for the first time, defined experimentally the extent to which, and mechanisms by which a specific biomolecule interacts with a range of well-defined, macroscopic surfaces. Here we propose a two-year research program aimed at adapting this quantitative experimental tool to the study of protein-surface interactions.