title: Rapid purification of native SecA from Escherichia coli: development of a new affinity chromatography procedure.
abstract: The SecA protein occupies a pivotal position in the public protein export pathway in Escherichia coli. The multifunctional SecA protein recognizes cytoplasmic factors associated with export including the presecretory protein and targets the complex to the inner membrane, where it acts in the early stages of protein translocation. The ability of SecA to bind ATP was the basis for the development of a novel, rapid purification scheme involving a single chromatographic step. Affinity chromatography was carried out on Red Sepharose CL-6B. The SecA present in crude extracts of E. coli binds strongly to this dye-ligand matrix, and active protein was purified to greater than 90% homogeneity. The protein isolated by this procedure retained the previously described ATPase and RNA-binding activities of SecA. This approach should permit the rapid purification of SecA homologs from a variety microorganisms.
authors.affiliation: Department of Microbiology and Immunology, Medical University of South Carolina, Charleston 29425-2230.
authors.forename: K B
authors.initials: KB
authors.lastname: Kiser
authors.forename: P
authors.initials: P
authors.lastname: Arnaud
authors.forename: M G
authors.initials: MG
authors.lastname: Schmidt
mesh_terms: D000251:Adenosine Triphosphatases / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods*; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D012685:Sepharose / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D000069816:SEC Translocation Channels; D012685:Sepharose; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins