Collaborative Research in Chemistry (CRC): Moderate Resolution Protein Structures by Chemical Crosslinking and Mass Spectrometry

罗德尼·K加州弗朗西斯科大学和加州圣巴巴拉大学的Guy、Bradford吉布森和欧文Kuntz因其在质谱法测定蛋白质结构方面的跨学科合作而得到特别项目办公室化学部的联合支持。他们将设计和合成交联剂，当它们在一定距离内时，这些交联剂将选择性地结合到氨基酸侧链对上。通过单独或组合使用各种交联剂，它们将能够化学连接相距5-20埃的残基。还将研究同位素标记的交联剂或掺入亲和纯化标签的交联剂。交联后，修饰的蛋白质将被消化，通过HPLC纯化，然后通过质谱分析。将开发计算技术来分析质谱数据并推导出蛋白质的三级结构。最初研究的蛋白质包括FGF-2和HIV整合酶。这种方法也将用于研究蛋白质与蛋白质或结构域与结构域之间的相互作用，确定蛋白质或蛋白质复合物的三级或三维结构是一个关键问题。这一质谱工具将为现有的核磁共振和单晶X射线衍射技术提供补充。我们的目标是开发一个高度自动化，高通量的系统，可以推断三级结构使用少量的蛋白质。化学合作研究（CRC）奖是给跨学科的科学家团队工作的问题，广泛的化学兴趣。这些奖项的重点是研究和培训的新合作模式。

Rodney K. Guy, Bradford Gibson and Irwin Kuntz of the University of California San Francisco and the University of California Santa Barbara are jointly supported by the Division of Chemistry, Special Projects Office, for their interdisciplinary collaboration on protein structure determinations by mass spectrometry. They will design and synthesize cross-linking agents that will selectively bind to pairs of amino acid sidechains when they are within a certain distance. By using a variety of crosslinking agents singly or in combination, they will be able to chemically connect residues that are 5-20 Angstroms apart. Isotopically labeled crosslinkers or crosslinkers that incorporate an affinity purification tag will also be investigated. After crosslinking, the modified proteins will be digested, purified by HPLC, then analyzed by mass spectrometry. Computational techniques will be developed to analyze the mass spectrometric data and derive the tertiary structure of the protein. Initial proteins to be studied include FGF-2 and HIV-integrase. This method will also be used to study protein-protein or domain-domain interactions.Determining the tertiary or three dimensional structure of proteins or protein complexes is a key problem. This mass spectrometric tool will provide a complement to existing techniques of nuclear magnetic resonance and single crystal x-ray diffraction. The goal is to develop a highly automated, high-throughput system that can infer tertiary structure using small amounts of protein.Collaborative Research in Chemistry (CRC) awards are given to interdisciplinary teams of scientists working on problems of broad chemical interest. The emphasis in these awards is on new collaborative modes of research and training.