IDBR: Closed-Loop Protein Crystallization

布兰迪斯大学的Fraden博士获得了一笔赠款，用于开发使蛋白质结晶的微流体工具。虽然蛋白质结晶的基本机制是由研究界商定的，但标准的结晶实践并不包括最佳地利用结晶背后的物理原理的方法和仪器。为了克服当前结晶技术的不足，Fraden博士将开发闭环蛋白质结晶器（CLC），该结晶器结合了微流体技术的高通量、精确度和低体积特性。CLC的概念基础是增加过饱和度直到检测到纳米尺度的晶核，然后动态地改变过饱和条件以控制晶体生长速率。微流控蛋白质结晶装置（CLC）由威尔斯阵列组成，以存储数百纳升体积的含有蛋白质溶液的液滴。该装置包括一个半透膜，允许蛋白质溶液的浓度变化，以及一个温度阶段，允许液滴在不同的威尔斯井经历不同的温度。CLC将结合动态光散射（DLS），用于检测成核和随后的晶体生长的开始。DLS的输出将用于闭环反馈模式，以控制结晶发生时刻的浓度和温度的热力学变量，这是控制结晶的最佳时刻。结构生物学领域致力于解决蛋白质的结构，其动机是相信？结构意味着功能。结构？功能关系对于包括新生物分子的新材料的仿生工程的基础知识是重要的。蛋白质结构通常通过X射线衍射获得，结晶是该过程中的必要步骤。构成人类基因组的30，000种蛋白质中有三分之一位于膜上，但只有一种人类膜蛋白的结构得到了解决。结晶是这一过程中的瓶颈，本项目的目标是通过开发闭环结晶器来消除这一瓶颈。此外，该项目所涉及的尖端芯片实验室仪器将为对美国经济和科学基础设施至关重要的技术部门的学生提供培训。

A grant has been awarded to Dr. Fraden at Brandeis University to develop microfluidic tools to crystallize proteins. Although the basic mechanism of protein crystallization is agreed upon by the research community, standard crystallization practices do not incorporate methods and instruments that optimally exploit the physical principles underlying crystallization. To overcome the deficiencies of current crystallization technologies Dr. Fraden will develop the Closed-Loop Protein Crystallizer (CLC), which incorporates the attributes of high-throughput, precision, and low volume that are characteristics of microfluidics. The conceptual underpinning of the CLC is to increase supersaturation until nanometer scale crystal nuclei are detected and then to dynamically vary the supersaturation conditions to control the rate of crystal growth.The microfluidic protein crystallization device (CLC) consists of an array of wells to store hundreds of nanoliter volume drops containing protein solution. The device incorporates a semi-permeable membrane that allows the concentration of the protein solution to be varied, as well as a temperature stage that allows drops in separate wells to experience different temperatures. The CLC will incorporate Dynamic Light Scattering (DLS) which will be used to detect the onset of nucleation and subsequent crystal growth. The output of the DLS will be used in a closed-loop feedback mode to control the thermodynamic variables of concentration and temperature at the moment of crystallogenesis, which is the optimal moment to control crystallization. The field of Structural Biology is dedicated to solving the structure of proteins motivated by the belief that ?structure implies function?. Structure ? function relationships are important for fundamental knowledge biomimetic engineering of new materials including new biomolecules. Protein structure is most commonly obtained through x-ray diffraction and crystallization is a necessary step in this process. One third of the 30,000 proteins that comprise the human genome reside in the membrane, yet only one human membrane protein has had its structure solved. Crystallization is the bottleneck in this process and the goal of this project is to eliminate this bottleneck by the development of the Closed Loop Crystallizer. Additionally, the cutting edge lab-on-a-chip instrumentation involved in this project will provide training to students in a technology sector vital to the US economy and scientific infrastructure.