Prelytic and lytic conformations of erythrocyte-associated Escherichia coli hemolysin.

Infection and Immunity
M Moayeri, R A Welch

Abstract

Flow cytometry was developed as a method to assess the conformation of erythrocyte-bound Escherichia coli hemolysin polypeptide (HlyA). Topology of membrane-associated hemolysin (HlyA(E)) was investigated by testing surface accessibility of HlyA regions in lytic and nonlytic bound states, using a panel of 12 anti-HlyA monoclonal antibodies (MAbs). Hemolysin associates nonlytically with erythrocytes at 0 to 2 degrees C. To test the hypothesis that the nonlytic HlyA(E) conformation at 0 to 2 degrees C differs from the lytic conformation at 23 degrees C, MAb epitope reactivity profiles at the two temperatures were compared by flow cytometry. Four MAbs have distinctly increased reactivity at 0 to 2 degrees C compared to 23 degrees C. HlyA requires HlyC-dependent acylation at lysine residues 563 and 689 for lytic function. Toxin with cysteine substitution mutations at each lysine (HlyA(K563C) and HlyA(K689C)) as well as the nonacylated form of hemolysin made in a HlyC-deficient strain were examined by flow cytometry at 0 to 2 and 23 degrees C. The three mutants bind erythrocytes at wild-type toxin levels, but there are conformational changes reflected by altered MAb epitope accessibility for six of the MAbs. To test further the surf...Continue Reading

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Citations

Jun 10, 2010·FEMS Microbiology Reviews·Irena LinhartováPeter Sebo
Jan 13, 2001·The Journal of Biological Chemistry·A L CortajarenaH Ostolaza
Sep 22, 2012·Immunopharmacology and Immunotoxicology·Nicole A AulikDarrel R McCaslin
Dec 20, 2005·Molecular Microbiology·Carlos BalsalobreSun Nyunt Wai
Aug 25, 2015·Biochimica Et Biophysica Acta·Laura C Ristow, Rodney A Welch
Jul 13, 2014·Analytical Biochemistry·Sabrina ThomasLutz Schmitt
Sep 2, 1999·Trends in Microbiology·E T LallyJ Korostoff
Jun 10, 1998·Microbiology and Molecular Biology Reviews : MMBR·P StanleyC Hughes

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