Quantitative Molecular Dynamics of Extremophile Metalloproteins -- Combining Experiment and Computation

这是SETI研究所、圣何塞州立大学、PCOM Georgia和乔治城大学之间的一个合作研究项目，旨在研究“极端微生物”的蛋白质。这些生物可以在极端环境中茁壮成长，例如低温（0°C，“嗜冷菌”）、高温（100°C，“超嗜热菌”）或高压（1100 atm，“嗜热菌”），以及其他极端的pH值、盐浓度和辐射。了解生物如何在极端条件下繁衍，对于指导在宇宙其他地方寻找生命非常重要。它还有许多实际应用，因为来自这些微生物的酶，“极端酶”，是非常有用的。它们在极端条件下的稳定性使得许多应用成为可能，例如冷水“生物洗涤剂”中的酶，用于更环保的纸张加工的高温“生物制浆”酶，以及用于食品加工的高压酶。尽管它们有许多应用，但导致极端酶稳定性的因素仍未完全了解。一些理论提出，这些蛋白质内部的分子运动决定了其稳定性和活性的范围。然而，对于不同条件下的极酶运动，目前还没有足够好的定量信息。在这个项目中，来自各种极端微生物的蛋白质将由圣何塞州立大学和乔治亚州PCOM大学的本科生生产。SETI的科学家将使用新型x射线技术来测量这些蛋白质的运动。研究结果将由乔治城大学的研究生进行理论计算测试。来自当地学校的代表性不足的高中生和本科生将参与实地考察和暑期研究项目。该项目旨在获得在一定温度和压力下，从嗜冷菌到超嗜热菌以及从嗜中菌到专性嗜湿菌的蛋白质原子运动的实验数据。利用核共振振动光谱（NRVS）进行的实验将提供rubredoxin和细胞色素P450活性位点的Fe原子和靶蛋白中间或外围的Te原子的运动信息。x射线衍射和核共振时域干涉法（NR-TDI）将量化这些蛋白质在多肽螺旋和环中的运动。这些数据将检验关于蛋白质动力学的两个假设。该项目将专门寻找“蛋白质玻璃转变”，这是在180-220 K范围内蛋白质灵活性的温度依赖性变化。该项目还将测试“对应状态”范式，该范式认为，极端微生物中的蛋白质已经被适应，在不同的环境条件下保持对每种生物有利的同等灵活性。最后，该项目旨在确定用于分子动力学（MD）模拟的力场和水模型的最佳组合，以重现和解释为一组蛋白质收集的实验数据。这项研究有望扩大对极端生物蛋白质分子运动的认识，以及在极端条件下促进生命的基本因素。该项目由分子和细胞生物科学部的分子生物物理集群资助。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This is a collaborative research project between the SETI Institute, San Jose State University, PCOM Georgia, and Georgetown University to study proteins from ‘extremophiles’. These are organisms that can thrive in extreme environments, such as low temperature (0°C, ‘psychrophiles’), high temperature (100°C, ‘hyperthermophiles’), or high pressure (1100 atm, ‘piezophiles’), as well as other extremes of pH, salt concentration, and radiation. Understanding how living things can flourish under extreme conditions is important for guiding the search for life elsewhere in the universe. It also has many practical applications, since the enzymes from these microorganisms, ‘extremozymes’, are extraordinarily useful. Their stability under extreme conditions has enabled numerous applications, such as enzymes included in cold-water ‘biodetergents’, high temperature ‘bio-pulping’ enzymes for more eco-friendly paper-processing, and high-pressure enzymes for food processing. Despite their numerous applications, the factors that lead to extremozyme stability are still not completely understood. Some theories propose that the molecular motion within these proteins determines the ranges of stability and activity. However, there is not enough good quantitative information about extremozyme motion under different conditions. For this project, proteins from various extremophiles will be produced by undergraduates at San Jose State and PCOM Georgia. The motion in these proteins will be measured by SETI scientists using novel x-ray techniques. The results will be tested against theoretical calculations by graduate students at Georgetown University. Underrepresented high school and undergraduate students from local schools will be involved in field trips and summer research projects.This project is undertaken to gain experimental data on atomic motion in proteins from psychrophiles to hyperthermophiles and from mesophiles to obligate piezophiles, under a range of temperatures and pressures. Experiments performed by using nuclear resonance vibrational spectroscopy (NRVS) will provide information on motion of Fe atoms at the active sites of rubredoxin and cytochrome P450 and Te atoms in the middle or on the periphery of the target proteins. X-ray diffraction and nuclear resonant time domain interferometry (NR-TDI) will quantify motion in polypeptide helices and loops of these proteins. The data will test two hypotheses concerning protein dynamics. The project will specifically look for ‘protein glass transitions’, which are proposed Temperature-dependent changes in protein flexibility over the range of 180-220 K. The project will also test the ‘corresponding states’ paradigm, which says that proteins in an extremophile have been adapted to maintain equivalent flexibility under the different environmental conditions favored for each organism. Finally, the project is aimed to determine the best combination of force fields and water models for molecular dynamics (MD) simulations to reproduce and interpret the experimental data collected for a set of proteins. This research is expected to expand the knowledge of molecular motion in extremophile proteins and the fundamental factors that contribute to life under extreme conditions. This project was funded by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.