CORE--SOLID PHASE PEPTIDE SYNTHESIS AND HPLC/CZE

该核心将合成和表征多肽，并开发新的基于高效液相和CZE的分析系统，以提高分辨率和效率。此外，该核心还将按要求进行圆二色(CD)实验。使用Merrifield自动SPPS，Core每年将总共合成大约20个多肽(长度为10到50个残基；线性、环状、磷酸化)。利用最新的制备性纯化技术，我们将提供10-100 mg的纯化多肽/蛋白质，用于高效液相色谱、FPLC离子交换层析和毛细管区带电泳(CZE)。必要时，将使用质谱仪(LSIMS)和Edman降解仪(C芯)进行表征。这些多肽将支持大多数项目将核心C将使用CZE。多肽将被用来进行功能和结构研究，以及产生特定的多克隆抗体。最后，我们打算利用这个核心的一些资源来开发新的分析技术和探索新的应用。这个核心将利用P.I.的S的经验(12%的努力)和他的技术助理以及他功能齐全的SPPS和表征实验室。实验室空间配备了引擎盖和其他小型设备。可用的主要仪器包括JEOL的HX-110型大规模集成电路质谱仪和60 MHz核磁共振，Beckman的P/ACE2050CZE，Beckman的三个多肽合成器，分析和制备HPLC和冻干机。P.I.将主持负责设定优先顺序的用户委员会。预计核心D将能够以先到先得的方式运作。在必须确定优先次序的情况下，将执行用户委员会大多数成员(主席除外)作出的决定。多肽的合成和纯化将由M.Odrowaz-Sypniewski进行(30%的工作)。D·柯比(15%的工作)将使用高效液相色谱、FPLC和CZE进行质量控制和方法开发。MS分析(15%的努力)将由A.Craig博士(15%的努力)进行。在征得有关P.I.的同意后，多余的多肽将免费分发给任何NIH赞助的研究人员，就像我们过去所做的那样。最近，人们对圆二色谱有很大的需求。我们已经负责该仪器十多年了，并将为该计划项目的调查人员提供分析。

This Core will synthesize and characterize peptides, and develop new HPLC- and CZE-based analytical systems to improve on resolution and efficiency. Additionally, this Core will perform circular dichroism (CD) experiments as requested. Using the Merrifield automated SPPS, Core will synthesize, each year, a total of approximately 20 peptides (10 to 50 residues in length; linear, cyclic, phosphorylated). Using the latest techniques for the preparative purification of peptide/proteins, we will provide 10 to 100 mg of purified peptides/proteins to HPLC, ion exchange chromatography on FPLC and capillary zone electrophoresis (CZE). Characterization will use mass spectroscopy (LSIMS) and Edman degradation (Core C) when necessary. These peptides will support most projects will Core C will use CZE. Peptides will be used to carry out functional and structural studies as well as to raise specific polyclonal antibodies. Finally, we intend to use some of the resources of this Core to develop new analytical techniques and explore new applications. This Core will take advantage of the P.,I.'s experience (12% effort) and that of his technical assistant as well as his fully functional laboratory for SPPS and characterization. Laboratory space equipped with hoods and other small equipment is available. Major instruments available include an LSI Mass Spectrometer Model HX-110 and a 60 MHz NMR from JEOL, a Beckman P/ACE model 2050 CZE, three peptide synthesizers from Beckman, analytical and preparative HPLCs and lyophilizers. The P.I. will chair the users' committee responsible for setting priorities. It is expected that Core D will be able to operate on a first come, first served basis. In cases where priority will have to be determined, the decision reached by the majority of the users' committee (excluding the chair) will be implemented. Synthesis and purification of peptides will be carried out by M. Odrowaz-Sypniewski (30% effort). Quality control and method development using HPLC, FPLC and CZE will be carried out by D. Kirby (15% effort). MS analyses (15% effort) will be carried out by Dr. A. Craig (15% effort). With the consent of the concerned P.I.s, excess peptides will be distributed free of charge to any NIH sponsored researcher as we have done in the past. Recently, there has been a significant need for circular dichroism spectroscopy. We have been responsible for the instrument for more than ten years and will provide analyses to investigators on this Program Project.