PROTEIN SEQUENCING FACILITY EXPANSION

来自威斯康星大学麦迪逊分校的十名用户正在申请一个 生物医学研究支持共享仪器补助金购买 设备，以扩大蛋白质微测序设施的新成立的 威斯康星州大学生物技术中心。 我们建议增加我们的 设施a）气相蛋白质微测序仪，以进行顺序 在亚纳摩尔规模下的蛋白质的Edman降解，B）用于测定蛋白质的HPLC。 在线分析所得PTH-氨基酸衍生物，c）HPLC， 在测序之前纯化蛋白质和肽，d）氨基酸 分析仪; e）必要的辅助设备。 测序亚纳摩尔量的蛋白质和肽的能力将 在研究难以纯化的蛋白质和获得 肽序列数据以确认DNA测序结果。 一个非常重要 令人兴奋的用途是克隆非丰富蛋白质的基因。 氨基酸 将获得序列信息并用于设计DNA 可以用作杂交探针以鉴定寡核苷酸， 基因文库中的目的基因。 本研究计画包括：1）酵素与蛋白质 调节遗传信息表达的中心（Burgess， Dahlberg，Reznikoff）; 2）参与蛋白质的结构和功能 钙通道（Kung）; 3）已知在人类疾病中缺乏的酶 （Pitot）; 4）小RNA病毒的结构和合成（Rueckert）; 5） 参与酰基辅酶A与ADP/ATP相互作用的蛋白质结构 载体（Shrago）; 6）维生素K依赖性羧化酶的酶学， 修饰凝血酶原（Suttie）的结构; 7）抗生素的控制 抗性（Weisblum）;和8）调节，细胞外输出和 溶细胞蛋白的结构（Welch）。 这个设施将给大量的教师谁是进行 NIH支持的研究增加了对一种非常强大的新技术的使用。 这项技术将大大有助于许多国家的成功。 这项研究，是必不可少的保持实力，并继续 这所大学的许多研究项目的进展。

Ten users from the University of Wisconsin-Madison are applying for a Biomedical Research Support Shared Instrumentation Grant to purchase equipment to expand a Protein Microsequencing Facility of the newly formed University of Wisconsin Biotechnology Center. We propose to add to our facility a) a gas-phase protein microsequencer to carry out sequential Edman degradation of proteins at a subnanomole scale, b) an HPLC for analyzing on line the resulting PTH-amino acid derivatives, c) an HPLC for purifying proteins and peptides prior to sequencing, d) an amino acid analyzer, and e) necessary accessory equipment. The ability to sequence subnanomole amounts of proteins and peptides will be invaluable in studies of difficult-to-purify proteins and in obtaining peptide sequence data to confirm DNA sequencing results. A very important and exciting use is to clone genes for non-abundant proteins. Amino acid sequence information will be obtained and used to design DNA oligonucleotides that can be used as a hybridization probe to identify the gene of interest in gene libraries. Projects proposed here include the study of: 1) enzymes and proteins central to regulating the expression of genetic information (Burgess, Dahlberg, Reznikoff); 2) structure and function of proteins involved in calcium channels (Kung); 3) enzymes known to be deficient in human disease (Pitot); 4) structure and synthesis of small RNA viruses (Rueckert); 5) structure of proteins involved in the interaction of acyl CoAs with ADP/ATP carrier (Shrago); 6) enzymology of vitamin K-dependent carboxylase and structure of modified prothrombin (Suttie); 7) control of antibiotic resistance (Weisblum); and 8) regulation, extracellular export and structure of cytolytic proteins (Welch). This facility will give a large number of faculty who are conducting NIH-supported research increased access to a very powerful new technology. This technology will contribute significantly to the success of much of this research and is essential for maintaining the strength and continued progress of many research programs at this University.