Expression and purification of soluble adenylyl cyclase from Escherichia coli

Methods Mol Biol. 2004;237:39-53. doi: 10.1385/1-59259-430-1:39.

Abstract

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.

Publication types

  • Research Support, U.S. Gov't, P.H.S.
  • Review

MeSH terms

  • Adenylyl Cyclases / genetics*
  • Adenylyl Cyclases / isolation & purification*
  • Adenylyl Cyclases / metabolism
  • Chromatography, Affinity / methods*
  • Escherichia coli / genetics*
  • Escherichia coli Proteins / genetics*
  • Escherichia coli Proteins / isolation & purification*
  • Escherichia coli Proteins / metabolism
  • Signal Transduction
  • Solubility

Substances

  • Escherichia coli Proteins
  • Adenylyl Cyclases