PROTEIN-PEPTIDE SEQUENCING FACILITY

这项建议的目的是建立一个“最先进的 用于蛋白质分析和表征的“设施”， 用自动微量测序技术测定肽 氨基酸组成，高效液相色谱 和DNA寡核苷酸合成。 我们的目标是建立 集中的蛋白质结构设施设在一个实验室。 要求的主要仪器是：（i）应用生物系统 在线脉冲液相蛋白质/肽测序仪 苯妥因衍生物的鉴定;（ii）应用 Biosystems Derivatizer氨基酸分析仪 氨基酸组成测定; iii）沃茨高 高效液相色谱系统 蛋白质、肽、苯硫基乙内酰脲的分析纯化 和寡核苷酸以及（iv）应用生物系统DNA合成仪 用于制备寡核苷酸。 这种仪器和 最近获得的肽合成器将为研究人员提供 有三种基本的方法：蛋白质测序， 肽合成和DNA合成。 该设施将确保 研究人员可以从蛋白质、信使RNA或基因开始， 从蛋白质中产生基因或从基因中产生蛋白质。 这种蛋白质结构仪器的可用性是 这对持续生产力和最佳进展至关重要， 阿尔伯特·爱因斯坦学院的广泛研究项目 医学正在解决生物学中的基本问题， 调控 目前，该机构拥有一个八岁的贝克曼 890 C旋转杯测序仪;虽然这种仪器仍然是 操作它是不够敏感或可靠的现代 测序应用。 本申请（a）证明需要 以提供最新的中央设施， （B）描述了以下方法的可行性： 安装、操作和维护此类设施，以及（c） 概述了一系列需要蛋白质测序的项目， 他们将立即使用该设施。

The objective of this proposal is to establish a "state of the art facility" for the analysis and characterization of proteins and peptides using automated microsequencing techniques, determination of amino acid composition, high performance liquid chromatography and DNA oligonucleotide synthesis. Our goal Is to establish a centralized protein structure facility housed in one laboratory. The major instrumentation requested is: (i) an Applied Biosystems Pulsed Liquid Phase Protein/Peptide Sequencer with on-line identification of phenylthlohydantoin derivatives; (ii) an Applied Biosystems Derivatizer Amino Acid Analyzer System for the determination of amino acid composition; iii) a Waters High Performance Liquid Chromatography System for preparative and analytical purification of proteins, peptides, phenylthiohydantoins and oligonucleotides and (iv) an Applied Biosystems DNA synthesizer for the preparation of oligonucleotides. This instrumentation and a recently acquired peptide synthesizer will provide investigators with three types of essential methodologies: protein sequencing, peptide synthesis and DNA synthesis. The facility will ensure that investigators can start at the protein, messenger RNA or gene levels to generate "genes from proteins" or "proteins from genes". The availability of this protein structure instrumentation is essential for continued productivity and optimal progress in the broad spectrum of research programs at Albert Einstein College of Medicine that are addressing fundamental problems in biological regulation. Presently, this Institution possesses one eight year old Beckman 890 C spinning-cup sequencer; although this instrument is still operational it is not sufficiently sensitive or reliable for modern sequencing applications. This application (a) documents the need for an up-dated centralized facility which will be available to our entire scientific community; (b) describes the feasibility of installing, operating and maintaining such a facility and (c) outlines a series of projects that require protein sequencing and which will make immediate use of the facility.