Ink-Jet System for Protein Crystallization

描述（由申请人提供）：在这个项目中，MicroFab将设计和制造一个基于喷墨的蛋白质晶体筛选平台（CrystalJet(tm)）的原型，重点是难以结晶但对细胞功能至关重要的膜蛋白。原型平台将执行高通量初始蛋白质结晶筛选，以确定后续扩大到衍射质量晶体的参数。基于计算和结构的药物开发的信息通常是通过结晶膜蛋白的x射线衍射研究获得的。然而，膜蛋白的结晶往往不能产生衍射质量的晶体。膜蛋白结晶的困难可归因于亚稳溶解度狭窄和纯化蛋白的稀缺。解决这些问题的方法是减少蛋白质的消耗，并通过使用较小的筛选体积来缩小晶体生长条件的采样范围。喷墨点胶是一种能够可靠地提供小体积的技术，从皮升到微升，后者通过吸积。CrystalJet（tm）筛选平台将在微批和蒸汽扩散模式下运行，以提供实验灵活性。我们还将开发一次性喷墨点胶机，以解决交叉污染问题。将开发一种dip-n- sip模式，用于在<1000 nL的体积下沉积蛋白质和匹配体积的结晶/沉淀溶液。笛卡尔流体处理系统可以处理体积更大的流体，例如油和井中沉淀液，可达1000 nL。该平台将包括一个板处理器和板封印机，用于操作和密封晶体筛选板，环境控制以保持温度、湿度和清洁空气，以及晶体观察和图像捕获系统。将使用蛋白质标准（如溶菌酶、葡萄糖异构酶、thaumatin）和膜蛋白进行一系列筛选实验，以验证CrystalJet（tm）平台的操作。CrystalJet平台是一种自动化工具，可以执行更多的蛋白质晶体学筛选实验，同时与传统筛选系统相比，使用的蛋白质要少得多。最终，该工具将能够更快、更有效地发现药物靶点，以帮助开发有效的药物来治疗疾病。

DESCRIPTION (provided by applicant): In this project, MicroFab will design and fabricate a prototype ink-jet based protein crystal screening platform (CrystalJet(tm)) with emphasis on membrane proteins that are difficult to crystallize, but critical to cellular function. The prototype platform will perform high-throughput initial protein crystallization screening to identify parameters for subsequent scale-up to diffraction quality crystals. Information for computational and structure based drug development is typically obtained through x-ray diffraction studies of crystallized membrane proteins. However, the crystallization of membrane proteins often fails to produce diffraction quality crystals. Difficulties in crystallizing membrane proteins can be attributed to narrow metastable solubility regions and scarcity of purified protein. A solution to these problems is to decrease protein consumption and narrow the sampling of crystal growth conditions by using smaller screening volumes. Ink-jet dispensing is a technology capable of reliably delivering small volumes ranging from picoliter to microliter, the latter by accretion. The CrystalJet(tm) screening platform will operate in microbatch and vapor diffusion modes to offer experimental flexibility. We will also develop a disposable ink-jet dispenser to address cross contamination issues. A dip-n- sip mode will be developed for the deposition of protein and matching volumes of crystallization/precipitant solution at volumes of <1000 nL. A Cartesian fluid handling system will manipulate larger fluid volumes of >1000 nL, such as oil and well precipitant solutions. The platform will include a plate handler and plate sealer to manipulate and seal crystal-screening plates, environmental control to maintain temperature, humidity and clean air, and a crystal observation and image capture system. A series of screening experiments will be performed using protein standards (e.g., lysozyme, glucose isomerase, thaumatin) and membrane proteins to validate the operation of the CrystalJet(tm) platform. The CrystalJet platform is an automated tool to perform a greater number of protein crystallography screening experiments, while using significantly less protein in comparison to conventional screening systems. Ultimately, this tool will enable faster and more efficient drug target discovery to aid in the development of effective drugs to treat disease.