The Gradient Foam Cement of the Sandcastle Worm as a Model for New Materials

ID：MPS/DMR/BMAT(7623)0906014 PI：Stewart，Russell ORG：犹他州大学标题：沙堡蠕虫的梯度泡沫水泥作为新材料的模型特长：沙堡蠕虫驻留在加利福尼亚州海岸潮间带的复合矿物壳中，用于物理保护。蠕虫以沙粒和贝壳碎屑的形式意外地收集矿物成分，并以建造石墙的方式用小块蛋白质胶将它们粘合在一起，形成一个管子。这种胶在冷海水中凝结在30 S以内，通过氧化交联硬化成坚韧的皮革状。对这种生物粘合剂的技术兴趣源于它在水下粘合各种矿物基质的能力。该粘合剂以胶体悬浮液的形式分泌，具有较低的初始粘度和界面张力，很容易在潮湿的基材上扩散，但它具有足够的粘聚力，在固化前不会分散到海洋中。此外，水性生物粘附剂显然取代了界面水，这是实现强大表面粘附性的先决条件。这些特性使P.calfornica粘合剂成为设计水性水下粘合剂的一种有价值的新范例。关于粘合剂的组成和生物加工，还有几个问题有待回答。举例来说，粘附性腺体包含至少两种不同类型的分泌颗粒。(1)胶粒类型之间的胶蛋白是如何分布的？(2)每种胶粒类型的胶蛋白是否在粘附腺的不同区域表达？(3)分泌过程中颗粒内容物的混合是否触发了固化和固化反应。已经设计了实验来分析胶粘剂的组成和固化前的生物处理。第一组实验(目标1)将通过基因表达分析提供粘附腺生理学的广泛概述。第二组实验(目标2)将使用目标1产生的基因和抗体探针，通过检查空间表达模式来研究粘附腺的功能划分。第三组实验(目标3)将采用强大的方法和现代质谱仪来分析新鲜分泌物粘合剂和单个分泌物颗粒的蛋白质含量。多肽将被蛋白质分解产生，并用微液相色谱和串联质谱仪(LC/MS/MS)进行分析。蛋白质组学分析将提供直接证据，证明在表达调查中发现的新胶蛋白存在于胶中。更广泛的影响：这些研究的结果将在解决重要的技术问题方面取得进展，例如使用可注射和生物兼容粘合剂的植入物中骨与骨或骨与金属的‘湿结合’，以及其他‘水下’粘合剂的应用。“以一种新颖而有效的方式实现了教学和科研的结合。大部分的粘附腺EST(表达序列标签)数据库将在分子生物工程本科实验室课程期间开发，在该课程中，学生将学习构建cDNA文库、纯化质粒、自动DNA测序、生物信息学资源，以及如何使用这些工具来研究生物过程。学生们发现，与罐装的实验室练习相比，这种体验相当独特。结果事先是未知的，学生们有一个真正的机会去发现新的生物技术资源。学生掌握他们正在研究的基因，进行生物信息学分析，并提出关于他们的基因在生物黏附中的潜在作用的假设。

ID: MPS/DMR/BMAT(7623) 0906014 PI: Stewart, Russell ORG: University of UtahTitle: The Gradient Foam Cement of the Sandcastle Worm as a Model for New MaterialsINTELLECTUAL MERIT: The sandcastle worm Phragmatopoma californica resides for physical protection in a composite mineral shell in the intertidal zone along the coast of California. The worm gathers the mineral component adventitiously as sand grains and bits of seashell hash which it bonds together into a tube with small dabs of a proteinaceous glue in the manner of building a stone wall. The glue sets within 30 s under cold seawater and hardens to a tough leathery consistency through oxidative crosslinking. Technological interest in this bioadhesive stems from its ability to bond a diverse range of mineral substrates under water. The adhesive is secreted as a colloidal suspension with low initial viscosity and interfacial tension that readily spreads on wet substrates, yet it is sufficiently cohesive that it does not disperse into the ocean before setting. Moreover, the water-borne bioadhesive apparently displaces interfacial water, a prerequisite for strong surface adhesion. These properties make the P. californica adhesive a valuable new paradigm for the design of water-borne, underwater adhesives. Several questions remain to be answered about the composition and biological processing of the adhesive. To illustrate, the adhesive gland contains at least two distinct types of secretory granules. (1) How are the glue proteins distributed between the granule types? (2) Are the glue proteins of each granule type expressed in distinct regions of the adhesive gland? (3) Does the mixing of the granule contents during secretion trigger the setting and curing reactions. Experiments have been devised to analyze the composition and biological processing of the adhesive before it sets. The first set of experiments (aim 1) will provide a broad overview of adhesive gland physiology through gene expression analysis. The second set of experiments (aim 2) will use the genetic and antibody probes resulting from aim 1 to investigate functional partitioning of the adhesive gland by examining spatial expression patterns. The third set of experiments (aim 3) will deploy powerful methods and modern mass spectrometry instrumentation to analyze the protein content of freshly secreted adhesive and of individual secretory granules. Peptides will be proteolytically generated and analyzed by micro-liquid chromatography and tandem mass spec (LC/MS/MS). Proteomic analysis will provide direct evidence that the new glue proteins discovered in the expression survey are present in the glue. BROADER IMPACTS: The results of these studies will make progress towards solving important technological issues such as 'wet bonding' of bone-to-bone or bone-to-metal in implants using injectable and biocompatible adhesives as well as other 'under water' adhesive applications." Integration of teaching and research is achieved in a novel and effective way. The majority of the P. californica adhesive gland EST (expressed sequence tags) database will be developed during an undergraduate laboratory course in Molecular Bioengineering where students learn about construction cDNA libraries, purifying plasmids, automated DNA sequencing, bioinformatics resources, and how these tools are used to investigate biological processes. Students find this experience quite unique compared to canned laboratory exercises. The results are not known before hand, and the students have an authentic opportunity to discover new biotechnological resources. Students take ownership of the genes they are investigating, pursue the bioinformatics analysis, and develop hypotheses about the potential role of their gene in the bioadhesive.