Beeler Jeff A, Tang Wei-Jen
Ben-May Institute for Cancer Research and Committee on Neurobiology, University of Chicago, IL, USA.
Methods Mol Biol. 2004;237:39-53. doi: 10.1385/1-59259-430-1:39.
This chapter outlines methods to purify soluble adenylyl cyclase (AC7) expressed in an Escherichia coli (E. coli) heterologous expression system. Guidelines are provided for constructing the expression plasmids, optimizing expression, culturing, and purifying the proteins. Purification requires two chromatographic steps. A histidine tag (H6) is incorporated into the expression vector and utilized for affinity purification on a Ni-NTA column. Subsequently, an anion exchange column is employed to further purify the protein.
本章概述了在大肠杆菌（E. coli）异源表达系统中表达的可溶性腺苷酸环化酶（AC7）的纯化方法。提供了构建表达质粒、优化表达、培养和纯化蛋白质的指导方针。纯化需要两个色谱步骤。将组氨酸标签（H6）掺入表达载体中，并用于在镍-氮三乙酸（Ni-NTA）柱上进行亲和纯化。随后，使用阴离子交换柱进一步纯化蛋白质。
