Raising antibodies against OprD, an outer membrane protein of Pseudomonas aeruginosa using translational fusions to MalE

OprD is an outer membrane porin of Pseudomonas aeruginosa that mediates upt ake of basic amino acids, peptides as well as carbapenem antibiotics. Polyc lonal antibodies were raised against the OprD porin by creating protein fus ions between the Escherichia coli maltose binding protein and four OprD fra gments. These were expressed in E. coli and shown to be exported to the per iplasm. The fusion proteins were purified by amylose affinity chromatograph y and used to immunize rabbits intramuscularly. We established that MalE fu sions to OprD fragments retain maltose and amylose binding activities in vi vo and in vitro, confirming proper folding of the MalE domain of hybrid pro teins. Furthermore, we demonstrate that this strategy can be used to obtain specific antibodies against bacterial outer membrane proteins (OMPs). (C) 2001 Elsevier Science B.V. All rights reserved.