Collaborative Research: Unravelling the Bacterium-Mineral Interface-Nanoscale Structures and Forces

EAR-0525297，LOWER，俄亥俄州立大学研究基金会EAR-0525340，Bickmore，Brigham Young University EAR-0525151，Beveridge，Guelph University of Guelph细菌是地球上最多产的生物群，就其地理范围和跨地质时期的寿命而言。大多数细菌通过在固体颗粒(如矿物)表面建立栖息地来生存，这些固体颗粒可能位于土壤、地下或水环境中。细菌通过力“线索”感知矿物表面的存在，这使细胞能够物理地感觉到另一个表面。虽然分子间和分子内的力量主导着微生物的生命，但它们看似微不足道的大小和长度尺度使它们很难研究。这项拟议的研究将通过分子建模、力测量和特征良好的二氧化硅矿物和细菌的显微图像的独特组合来开始阐明这个问题，这些矿物和细菌已经被转基因，以产生特定的细胞壁大分子。首先，将用原子力滴定测量确定层状硅酸盐和硅酸盐晶体(可能也包括细菌表面)特定表面上官能团的自然分布、密度和酸碱反应活性。这些测量将用一种新的方法来解释，该方法用于预测单个官能团的酸碱反应活性，该方法涉及从头计算结构。其次，原子力显微镜将用于测量野生型和突变型铜绿假单胞菌和硅酸盐矿物之间的分子间力。这些力测量将根据先前获得的矿物表面酸碱反应的分子尺度模型来解释。这一提议的智力价值在于，将分子模型与力测量相结合，将允许以前所未有的方式观察活着的细菌和就地矿物表面之间的界面上存在的基本力量或线索。这一目标将通过我们的跨学科研究团队的共同努力来实现，其中包括专门从事地球化学、矿物学和分子建模(Bickmore和Lewis)、地球微生物学和纳米科学(LOWER和Beveridge)以及物理力定律(达彻和伊斯拉赫维利)的科学家。这些实验中使用的微生物铜绿假单胞菌是一种典型的革兰氏阴性细菌。它普遍存在于水、土壤和地下环境中，在这些环境中，它生活在矿物或其他颗粒的表面。它也是动植物上的一种常见细菌物种，在植物和动物中，它经常起到机会性病原体的作用。这些实验中使用的矿物包括硅酸盐，这是地球上最常见的无机相。探测铜绿假单胞菌和硅酸盐矿物之间的界面将具有更广泛的影响和社会效益，可应用于从含水层中微生物的运输到固体基质上生物膜的形成等一系列问题。此外，这项建议将影响一些研究生和本科生的生活。这些学生将接受生物化学、微生物学、地球化学和矿物学的交叉培训，他们还将获得扫描探针显微镜、激光扫描显微镜和透射电子显微镜等最先进仪器的经验。最后，这项提案将提供资金，支持旨在让年轻学生接触生物科学和物理科学之间相互作用的K-6推广计划。

EAR-0525297, Lower, Ohio State University Research FoundationEAR-0525340, Bickmore, Brigham Young UniversityEAR-0525151, Beveridge, University of Guelph Bacteria are the most prolific group of organisms on the Earth in terms of their geographic extent as well as their longevity across geologic time. Most bacteria live by creating habitats on the surface of solid particles such as minerals, which may be located in soil, subsurface, or aquatic environments. Bacteria perceive the presence of mineral surfaces through force "cues", which allow a cell to physically feel another surface. While inter- and intra-molecular forces dominate the lives of microorganisms, their seemingly infinitesimal magnitude and length-scale have made them difficult to study. This proposed research will begin to shed light onto this problem though a unique combination of molecular modeling, force measurements, and microscopic images of well-characterized silica minerals and bacteria that have been genetically modified to produce specific cell wall macromolecules. First, the natural distribution, density, and acid-base reactivity of functional groups on specific faces of phyllosilicate and silicate crystals (and possibly also on the surface of a bacterium) will be determined with atomic force titration measurements. These measurements will be interpreted with a new method for predicting acid-base reactivity of individual functional groups that involves ab initio structure calculations. Second, atomic force microscopy will be used to measure intermolecular forces between wild-type and mutant strains of Pseudomonas aeruginosa and silicate minerals. These force measurements will be interpreted in light of the previously obtained molecular-scale models of the mineral surface acid-base reactivities. The intellectual merit of this proposal is that the combination of molecular models with force measurements will allow an unprecedented view of the fundamental forces or cues that exist at the interface between a living bacterium and mineral surface in situ. This goal will be accomplished through the collective efforts of our interdisciplinary research team that includes scientists specializing in geochemistry, mineralogy and molecular modeling (Bickmore and Lewis), geomicrobiology and nanoscience (Lower and Beveridge), and physical force laws (Dutcher and Israelachvili). The microorganism used in these experiments, P. aeruginosa, is a model Gram negative bacterium. It is ubiquitous in water, soil, and subsurface environments where it lives on the surface of minerals or other particles. It is also a common bacteria species on plants and animals, where it often functions as an opportunistic pathogen. The minerals used in these experiments include silicates, which are the most common inorganic phases on Earth. Probing the interface between P. aeruginosa and silicate minerals will have broader implications and societal benefits that can be applied to issues ranging from the transport of microorganisms in aquifers to the formation of biofilms on solid substrates. Further, this proposal will impact the lives of a number of graduate students and undergraduates. These students will be cross trained in biochemistry, microbiology, geochemistry, and mineralogy, and they will also gain experience with state-of-the-art instruments such as scanning probe microscopes, laser scanning microscopy, and transmission electron microscopy. Finally, this proposal will provide funds to support K-6 outreach programs that are designed to expose young students to the interplay between the biological and physical sciences.