Expression, purification and preliminary crystallographic analysis of bacterial transmembrane protein EfeU for iron import

Abstract

The bacterial Efe system functions as an importer of free Fe²⁺ into cells independently of iron-chelating compounds such as siderophores and consisted of iron-binding protein EfeO, peroxidase EfeB, and transmembrane permease EfeU. While we and other researchers reported crystal structures of EfeO and EfeB, that of EfeU remains undetermined. In this study, we constructed expression system of EfeU derived from Escherichia coli, selected E. coli Rosetta-gami 2 (DE3) as an expression host, and succeeded in purification of the proteins which were indicated to form an oligomer by blue native PAGE. We obtained preliminary data of the X-ray crystallography, suggesting that expression and purification methods we established in this study enable structural analysis of the bacterial Efe system.

Keywords: Crystallization; Efe; EfeU; Escherichia coli; Iron importer; Siderophore-independent.