Peptide Synthesis and Purification Core Facility

A.局该设施将为主要描述的研究提供合成肽 项目2、3和4。该核心由Anantharamaiah博士指导，Palgunachari博士将协助 这个设施的日常运作这一核心设施的生产力很高， 广泛支持该计划项目，因此，该计划的重要组成部分 项目应用。 B。仪器仪表。该设施拥有三个蛋白质技术自动肽 合成器和一个Advanced ChemTech（ACT 90）肽合成器。而蛋白质 技术肽合成器可以提供少量的肽（mg量）， 更短的持续时间，因为在氨基上添加了十倍过量的进入氨基酸， 酸性树脂，ACT可以提供10 g量的肽，但由于一个氨基酸， 在合成的每个阶段确定反应完成后，每天加入酸。 由于ACT 90中使用的起始树脂更多，我们避免添加超过所需过量的 在每一阶段都使用氨基酸衍生物，以节省氨基酸衍生物的成本。为 核心设施拥有三台Beckman分析型HPLC和两台 制备型HPLC单元（Beckman和Varian）。除此之外，核心设施还利用BioRad 连接至Michel Muller中压液相色谱仪的FPLC系统 （MPLC）玻璃柱。这些列的大小不同，最小的（分析）列包含 12 g固定相（C-4或C-18硅胶）和含有800 g “自制”反相固定相（C-4和C-18反相硅胶）。我们 通常用各种链长的脂肪酸烷基化硅胶。我们一次生产2公斤， 国产色谱柱固定相为4kg。这提供了更高的能力，以净化大 大量的合成肽。这些柱也用于纯化重组蛋白。一 典型的程序将在方法部分中描述。

A. Administration. This facility will provide synthetic peptides for studies described mainly in projects 2, 3 and 4. This core is directed by Dr. Anantharamaiah, and Dr. Palgunachari will assist in the day-to-day operation of this facility. This core facility has been highly productive and has supported this program project extensively and therefore, an important component of this program project application. B. Instrumentation. The facility possesses three Protein Technology automatic peptide synthesizers and one Advanced ChemTech (ACT 90) peptide synthesizer. While the Protein Technology peptide synthesizers can provide small amounts of the peptides (in mg quantities) in a shorter duration due to the addition of ten times excess of incoming amino acids on to the amino acid-resin, the ACT can provide 10g quantities of peptides but the process is slow since one amino acid is added per day after the completion of reaction is ascertained at each stage of the synthesis. Since the starting resin used in ACT 90 is more, we avoid adding more than the required excess of amino acid derivatives at each stage in order to save costs on amino acid derivatives. For the purification of peptides the core facility possesses three Beckman analytical HPLC and two preparative HPLC units (Beckman and Varian). In addition to this, the core facility utilizes BioRad FPLC systems that are connected to Michel Muller Medium Pressure Liquid Chromatography (MPLC) glass columns. These columns are of various sizes with the smallest (analytical) containing 12g of the stationary phase (C-4 or C-18 silica gel) and the biggest column containing 800g of "home made" reverse phase stationary phase (C-4 and C-18 Reversed Phase silica gels). We routinely alkylate Silica Gel with various chain-lengths of fatty acids. At a time we produce 2 kg to 4kg of the stationary phase for home made columns. This provides a higher capacity to purify large quantities of synthetic peptides. These columns are also used to purify recombinant proteins. A typical procedure will be described in the Methods Section.