Oriented Protein Microarray for Functional Studies

描述（由申请人提供）： 拟议的研究旨在开发蛋白质微阵列作为蛋白质组学市场的上级技术导向和可行的产品。在第一阶段的研究中，PI和同事们成功地证明了制造具有高选择性和特异性的蛋白质微阵列的可行性，固定化蛋白质分子的方向均匀且可控，并且蛋白质处于天然状态，易于被溶液中的目标接近。更重要的是，初步研究不仅建立了这种化学在选择性蛋白质固定而不需要预纯化方面的优势，而且还建立了确保天然状态活性的取向控制的必要性。后者被证明的观察，在表面上的定向磺基转移酶的酶活性是相同的溶液相酶，而随机取向的酶在表面上显示出小于20%的活性。在第二阶段的资助下，我们建议（1）建立在表面上保持天然活性的定向蛋白质的普遍性;（2）开发固定化蛋白质的芯片上重折叠方法;（3）进一步开发商业化的表面涂层技术。第二阶段研究的成功将导致推出一种用于蛋白质微阵列的上级表面涂层技术，该技术具有比蛋白质微阵列市场中现有技术更高的性能和潜在更低的成本，这对于未来的大规模药物筛选和疾病诊断非常重要。

DESCRIPTION (provided by applicant): The proposed research is aimed at developing protein microarrays as a superior technology oriented and viable product for the proteomics market. During Phase-I research, the PI and coworkers have successfully demonstrated the feasibility of fabricating protein microarrays with high selectivity and specificity, with uniform and controllable orientation of immobilized protein molecules, and with proteins in native states and easily accessible by targets in the solution. More importantly, preliminary research established not only the advantage of this chemistry in terms of selective protein immobilization without the need of pre-purification but also the necessity of orientational control in ensuring activity of the native state. The latter was demonstrated by the observation that enzymatic activity of oriented sulfotransferases on the surface was the same as that of solution phase enzymes, while randomly oriented enzymes on the surface showed less than 20% of the activity. Under Phase II funding, we propose (1) to establish the generality of oriented proteins on the surface in maintaining native activity; (2) to develop methods for on-chip refolding of immobilized proteins; (3) to further develop surface coating technology for commercialization. The success of Phase-II research will lead to the launch of a superior surface coating technology for protein microarrays with higher performance at potentially lower cost than current technology in the protein microarray market, which is very important for future large scale drug screening and disease diagnosis.