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

就其地理范围和寿命而言，细菌是地球上最多产的生物群体。大多数细菌通过在固体颗粒（如矿物质）表面创造栖息地来生存，这些颗粒可能位于土壤、地下或水生环境中。细菌通过力“线索”感知矿物表面的存在，这使得细胞能够物理地感受到另一个表面。虽然分子间和分子内的作用力支配着微生物的生命，但它们看似无穷小的大小和长度尺度使它们难以研究。这项拟议的研究将通过分子建模、力测量和经过基因改造以产生特定细胞壁大分子的二氧化硅矿物和细菌的显微图像的独特组合，开始阐明这一问题。首先，用原子力滴定法测定层状硅酸盐和硅酸盐晶体特定表面（也可能是细菌表面）官能团的自然分布、密度和酸碱反应性。这些测量结果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铜绿假单胞菌野生型和突变株与硅酸盐矿物之间的分子间力。这些力的测量结果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。这一建议的智力价值在于，分子模型与力测量的结合将使我们能够前所未有地了解存在于活细菌和矿物表面之间的界面上的基本力或线索。这一目标将通过我们跨学科研究团队的共同努力来实现，该团队包括专门研究地球化学、矿物学和分子模型（Bickmore和Lewis）、地球微生物学和纳米科学（Lower和Beveridge）以及物理力定律（Dutcher和Israelachvili）的科学家。在这些实验中使用的微生物，铜绿假单胞菌，是一种模式革兰氏阴性菌。它在水、土壤和地下环境中无处不在，它生活在矿物质或其他颗粒的表面。它也是植物和动物上常见的细菌物种，在那里它经常作为机会性病原体发挥作用。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。探索铜绿假单胞菌和硅酸盐矿物之间的界面将具有更广泛的意义和社会效益，可以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问题。此外，这项提案将影响许多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、地球化学和矿物学方面的交叉培训，他们还将获得使用最先进仪器的经验，如扫描探针显微镜、激光扫描显微镜和透射电子显微镜。最后，该提案将为K-6扩展项目提供资金支持，这些项目旨在让年轻学生接触到生物和物理科学之间的相互作用。

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.