Purification of outer membrane iron transport receptors from Escherichia coli by fast protein liquid chromatography: FepA and FecA.
Fast protein liquid chromatography (FPLC) with DEAE-Sepharose Fast Flow, PBE-94 and Q-Sepharose Fast Flow columns are applied to the purification of the ferric enterobactin protein receptor (FepA). The apparent single band of FepA on SDS-PAGE is isolated and purified into two proteins with very similar molecular weights. The two proteins are identified to be FepA and ferric citrate protein receptor (FecA) by N-terminus amino acid determination and a computer search with the Gene Bank file. The assay of binding activities of these proteins shows that both FepA and FecA bind ferric enterobactin, with the former having about double the activity of the latter. Competition studies shows that Fe-MECAM is competitively bound to both proteins and that ferric parabactin only slightly competes with [55Fe]ferric enterobactin. It is found that ferrichrome A has no effect on the binding of the receptor proteins with ferric enterobactin.
Zhou
X H
XH
Department of Chemistry and Biochemistry, University of Oklahoma, Norman 73019.
van der Helm
D
D
Adjimani
J
J
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D002853:Chromatography, Liquid; D002951:Citrates / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004758:Enterobactin / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins; D007501:Iron / Q000378:metabolism*; D008969:Molecular Sequence Data; D011956:Receptors, Cell Surface; D011955:Receptors, Drug / Q000302:isolation & purification* / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D002951:Citrates; D029968:Escherichia coli Proteins; C028589:FecA protein, E coli; D011956:Receptors, Cell Surface; D011955:Receptors, Drug; C029489:enterobactin receptor; D004758:Enterobactin; D007501:Iron