The application of 0.1 M quadrol to the microsequence of proteins and the sequence of tryptic peptides.
The application of 0.1 M quadrol to the microsequence of proteins and the sequence of tryptic peptides.
复制标题
0.1 M 乙二胺在蛋白质微序列和胰蛋白酶肽序列中的应用。
作者：
A. W. Brauer;M. N. Margolies;E. Haber
为了将自动Edman降解扩展到纳摩尔量的蛋白质，Edman和Begg（Edman，P.，和Begg，G.（1967），Eur. J. Biochem.1，80）进行修饰以允许在不存在载体蛋白的情况下长时间降解。通过使用具有有限的、组合的苯-乙酸乙酯溶剂提取的0.1 M Quadrol水溶液程序，以及改变七氟丁酸的递送系统，可以从在7 nmol肌红蛋白上运行的测序仪中回收和鉴定前30个氨基酸残基。对于3 nmol的肌红蛋白，可以确定20个步骤。用气相色谱法和聚酰胺薄层色谱法对PTH-氨基酸进行了鉴定。在不使用载体蛋白的情况下，杯子可以防止机械损失（Niall，H. D、雅各布斯，J.W.，货车Rietshoten，J.，和Tregear，G. W.（1974），FEBS Lett. 41，62），使用该程序的重复产率为93- 96%。同样的程序已成功地应用于14个或更多个残基的肽，用或不用Braunitzer试剂修饰，并应用于许多较大的肽和蛋白质，包括兔抗体重链的216个残基片段，其中在25 nmol上完成了35个步骤的序列。
In an effort to extend automated Edman degradation to nanomole quantities of protein, the method of sequenator analysis described by Edman and Begg (Edman, P., and Begg, G. (1967), Eur. J. Biochem. 1, 80) was modified to permit long degradations in the absence of carrier proteins. By using an aqueous 0.1 M Quadrol program with limited, combined benezene-ethyl acetate solvent extractions, as well as a change in the delivery system for heptafluorobutyric acid, it was possible to recover and identify the first 30 amino acid residues from a sequenator run on 7 nmol of myoglobin. For 3 nmol of myoglobin, 20 steps could be identified. PTH-amino acids were identified by gas-liquid chromatography and thin-layer chromatography on polyamide sheets. Without using a carrier protein the cup to prevent mechanical losses (Niall, H. D., Jacobs, J. W., Van Rietshoten, J., and Tregear, G. W. (1974), FEBS Lett. 41, 62), the repetitive yield using this program was 93-96%. The same program has been applied successfully to peptides of 14 or more residues with or without modification by Braunitzer's reagent and to a number of larger peptides and proteins including a 216 residue segment of rabbit antibody heavy chain in which a sequence of 35 steps was accomplished on 25 nmol.