Synthesis and properties of a bacterial bioadhesive

描述（由申请人提供）：细菌通常利用多糖作为粘附结构附着在表面，形成生物膜，并感染宿主细胞。此外，多糖在人类活动的许多领域作为生物粘合剂，包括牙科和外科粘合剂，具有很强的前景。月牙根杆菌合成了一种叫做固化剂的多糖，这种多糖表现出令人印象深刻的粘附力。与大多数商用粘合剂不同，固定式粘合剂在淡水和海洋环境中都能紧紧粘附在各种表面上。这种特性对人体的医学应用至关重要。本研究的总体目标是使用从遗传学到生物物理学的多学科方法来研究固持特性的化学和生物物理基础，并了解固持特性是如何通过细胞外DNA （eDNA）的去乙酰化和抑制来调节的。该项目有三个具体目标。第一个目的是确定固化剂粘附性的生物物理基础。原子力显微镜（AFM）将用于系统地研究表面粗糙度、剪切应力、表面组成、离子强度和pH值对固结剂粘附的影响，以便更好地了解固结剂的粘附机理和粘附控制。第二个目的是确定去乙酰化在固相锚定和粘接性能中的作用。钉子户多糖脱乙酰酶突变导致钉子户在溶液中释放不粘附的钉子户。从正常细胞和脱乙酰酶突变细胞中可以确定钉子蟹多糖的组成和结构。AFM力谱和高分辨率荧光显微镜将用于确定脱乙酰化对持荷剂粘附性和内聚性的作用。生化实验将用于研究去乙酰化在固定固定物到细胞中的作用。最后，对海洋生物的固持进行类似的研究，将为在人体盐水环境中的潜在应用提供更好的生物材料。第三个目的是确定最近发现的eDNA抑制hold - fast粘附机制的生物学基础。将研究毒素-抗毒素系统在程序性细胞死亡产生eDNA中的作用，并确定hold - fast抑制序列特异性的基础。AFM压痕研究以及同时AFM成像和拉曼散射光谱将用于确定特定DNA如何改变固定物的结构和结构特性。所提出的研究结果将提供对holdfast令人印象深刻的粘合特性的基本机制和这些特性的调节的见解，为holdfast作为生物粘合剂的未来发展铺平道路。此外，这些研究的结果将提供见解

DESCRIPTION (provided by applicant): Bacteria often utilize polysaccharides as adhesive structures to attach to surfaces, to form biofilms, and to infect host cells. In addition, polysaccharides hold strong promise as biological adhesives in many areas of human activity, including as dental and surgical adhesives. The bacterium Caulobacter crescentus synthesizes a polysaccharide called the holdfast that exhibits and impressive adhesive force. Contrary to most commercial adhesives, holdfasts adhere tightly to a variety of surfaces in both freshwater and marine environments. Such a property is critical for medical applications in the human body. The general goal of this research is to use a multidisciplinary approach ranging from genetics to biophysics to study the chemical and biophysical basis for holdfast properties and to understand how holdfast properties are modulated by deacetylation and inhibition by extracellular DNA (eDNA). The project has three specific aims. The first aim is to determine the biophysical basis for holdfast adhesiveness. Atomic force microscopy (AFM) will be used to systematically study the influence of surface roughness, shear stress, surface composition, ionic strength, and pH on holdfast adhesion in order to provide a better understanding of the mechanism of holdfast adhesion and adhesion control. The second aim is to determine the role of deacetylation in holdfast anchoring and adhesive properties. A holdfast polysaccharide deacetylase mutant causes the release of non-adherent holdfast in solution. The composition and structure of the holdfast polysaccharide will be determined from normal and deacetylase mutant cells. AFM force spectroscopy and high-resolution fluorescence microscopy will be used to determine the role of deacetylation on holdfast adhesiveness and cohesiveness. Biochemical experiments will be used to study the role of deacetylation in anchoring the holdfast to the cell. Finally, similar studies of the holdfast of marine species will provide better biomaterials for potential applications in the saline environment of the human body. The third aim is to determine the biological basis for the recently discovered mechanism of eDNA inhibition of holdfast adherence. The role of a toxin-antitoxin system in the production of eDNA by programmed cell death will be studied and the basis for the sequence specificity of holdfast inhibition will be determined. AFM indentation studies and simultaneous AFM imaging and Raman scattering spectroscopy of holdfasts bound or not to eDNA will be used to determine how specific DNA alters the structure and structural properties of the holdfast. Results from the proposed studies will provide insight into the basic mechanisms for the impressive adhesive properties of the holdfast and modulation of these properties, paving the way for the future development of the holdfast as a biological adhesive. In addition, results of these studies will provide insights into the mechanism of polysaccharide adhesiveness in general, as well as for strategies to inhibit polysaccharide adhesion, for example during infection by pathogens.