CORE--SOLID PHASE PEPTIDE SYNTHESIS AND HPLC/CZE

该核心将合成和表征肽，并开发新的基于HPLC和CZE的分析系统，以提高分辨率和效率。此外，该核心将根据要求进行圆二色性（CD）实验。使用梅里菲尔德自动化SPPS，Core每年将合成总计约20种肽（长度为10至50个残基;线性、环状、磷酸化）。使用肽/蛋白质制备纯化的最新技术，我们将提供10至100 mg纯化肽/蛋白质，用于HPLC，FPLC离子交换色谱和毛细管区带电泳（CZE）。必要时，将使用质谱（LSIMS）和Edman降解（核心C）进行表征。这些肽将支持核心C将使用CZE的大多数项目。肽将用于进行功能和结构研究以及产生特异性多克隆抗体。最后，我们打算利用该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.的经验（12%的努力）和他的技术助理，以及他的全功能实验室的SPPS和表征。实验室空间配备了通风橱和其他小型设备。可用的主要仪器包括来自JEOL的HX-110型LSI质谱仪和60 MHz NMR、Beckman P/ACE 2050 CZE型、来自Beckman的三种肽合成仪、分析型和制备型HPLC以及冻干机。私家侦探将担任负责确定优先次序的用户委员会主席。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，将执行大多数用户委员会（不包括主席）作出的决定。肽的合成和纯化将由M. Odrowaz-Sypniewski（30%的努力）。采用HPLC、FPLC和CZE的质量控制和方法开发将由D.科比（15%的努力）。MS分析（15%工作量）将由Dr. A.克雷格（15%的努力）。在征得相关PI的同意后，多余的肽将免费分发给任何NIH资助的研究人员，就像我们过去所做的那样。近年来，人们对圆二色光谱学有着极大的需求。我们已经负责该仪器十多年了，并将为该计划项目的研究人员提供分析。

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.