Langelier Marie-France, Planck Jamie L, Servent Kristin M, Pascal John M
Department of Biochemistry, Molecular Biology and Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, USA.
Methods Mol Biol. 2011;780:209-26. doi: 10.1007/978-1-61779-270-0_13.
A general method to express and purify full-length human poly(ADP-ribose) polymerase-1 (PARP-1), individual PARP-1 domains, and groups of PARP-1 domains from Escherichia coli cells is described. The procedure allows for robust production of highly pure PARP-1 that is free of DNA contamination and well-suited for biochemical experiments and for structural and biophysical analysis. Two biochemical assays for monitoring PARP-1 automodification activity are presented that can be used to evaluate purified PARP-1, combinations of PARP-1 domains, or PARP-1 mutants.
本文描述了一种从大肠杆菌细胞中表达和纯化全长人聚（ADP - 核糖）聚合酶 -1（PARP -1）、PARP -1单个结构域以及PARP -1结构域组合的通用方法。该方法能够高效生产高度纯净且无DNA污染的PARP -1，非常适合用于生化实验以及结构和生物物理分析。文中还介绍了两种用于监测PARP -1自身修饰活性的生化检测方法，可用于评估纯化后的PARP -1、PARP -1结构域组合或PARP -1突变体。
