PMID: 7517392Jul 1, 1994Paper

Genetic insertion and exposure of a reporter epitope in the ferrichrome-iron receptor of Escherichia coli K-12

Journal of Bacteriology
G S MoeckJ W Coulton

Abstract

The ferrichrome-iron receptor of Escherichia coli K-12 is FhuA (M(r), 78,992), the first component of an energy-dependent, high-affinity iron uptake pathway. FhuA is also the cognate receptor for bacteriophages T5, T1, phi 80, and UC-1, for colicin M and microcin 25, and for albomycin. To probe the topological organization of FhuA which enables recognition of these different ligands, we generated a library of 16 insertion mutations within the fhuA gene. Each insertion spliced a 13-amino-acid antigenic determinant (the C3 epitope of poliovirus) at a different position within FhuA. Immunoblotting of outer membranes with anti-FhuA and anti-C3 antibodies indicated that 15 of 16 FhuA.C3 proteins were present in the outer membrane in amounts similar to that observed for plasmid-encoded wild-type FhuA. One chimeric protein with the C3 epitope inserted after amino acid 440 of FhuA was present in the outer membrane in greatly reduced amounts. Strains overexpressing FhuA.C3 proteins were subjected to flow cytometric analysis using anti-FhuA monoclonal antibodies. Such analysis showed that (i) the chimeric proteins were properly localized and (ii) the wild-type FhuA protein structure had not been grossly altered by insertion of the C3 epi...Continue Reading

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Citations

Oct 1, 2014·Proceedings of the National Academy of Sciences of the United States of America·Anna KonovalovaThomas J Silhavy
Sep 16, 2015·Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences·Anna Konovalova, Thomas J Silhavy
Nov 25, 1998·Protein Science : a Publication of the Protein Society·K DiederichsJ Breed
Feb 3, 1999·Annual Review of Genetics·J W Jarvik, C A Telmer
Jun 13, 2000·Protein Science : a Publication of the Protein Society·A D FergusonW Welte
May 23, 1997·The Journal of Biological Chemistry·R SrikumarJ W Coulton
Apr 18, 2007·Infection and Immunity·Amanda J DeRocco, Cynthia Nau Cornelissen
Jul 3, 2004·Journal of Bacteriology·Franziska Endriss, Volkmar Braun
Feb 24, 2004·Infection and Immunity·Mary Kate Yost-Daljev, Cynthia Nau Cornelissen
Aug 31, 2010·Applied and Environmental Microbiology·María Fernanda PomaresPaula A Vincent
Mar 19, 2008·Infection and Immunity·Jennifer M Noto, Cynthia Nau Cornelissen
Sep 28, 1998·Journal of Bacteriology·C J LazdunskiH Bénédetti
Jun 1, 1995·Journal of Bacteriology·R A Salomón, R N Farías

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