Use of a bacteriophage-encoded glycanase enzyme in the generation of lipopolysaccharide O side chain deficient mutants of Escherichia coli O9:K30 and Klebsiella O1:K20: role of O and K antigens in resistance to complement-mediated serum killing.

Canadian Journal of Microbiology
K L McCallumC Whitfield

Abstract

Coliphage K30 lysates contain free and phage-associated forms of a bacteriophage-encoded capsule depolymerase (glycanase) enzyme, active against the serotype K30 capsular polysaccharide of Escherichia coli. The free glycanase has been purified to apparent homogeneity. The molecular weight of the enzyme was estimated at 450,000, and when heated in SDS at 100 degrees C, the enzyme dissociated into two subunits of 90,000 and 52,000. The glycanase enzyme was used as a reagent to reversibly degrade the capsular layers on cells of Escherichia coli O9:K30 and Klebsiella O1:K20. This treatment rendered these bacteria sensitive to their respective lipopolysaccharide-specific bacteriophages, coliphage O9-1 and Klebsiella phage O1-3. This novel approach facilitated isolation of lipopolysaccharide O antigen side chain deficient mutants which retained the ability to synthesize the capsule. The response of defined mutants, O+:K-, O-:K+, and O-:K-, to exposure to nonimmune rabbit serum was measured. Results showed that the primary barrier against complement-mediated serum killing in both Escherichia coli O9:K30 and Klebsiella O1:K20 was the O antigen side chains of the lipopolysaccharide molecules. In both strains, the capsule played no role ...Continue Reading

Citations

Apr 16, 2002·Infection and Immunity·Linda J McKerral, Reggie Y C Lo
Apr 28, 2006·Proceedings of the National Academy of Sciences of the United States of America·Michael WackerMarkus Aebi
Mar 13, 2014·FEMS Microbiology Letters·Helen Miajlovic, Stephen G Smith
Nov 18, 2015·PloS One·Camila Figueiredo PinzanMaria Cristina Roque-Barreira
Mar 25, 2017·Applied Microbiology and Biotechnology·Agnieszka LatkaZuzanna Drulis-Kawa
Aug 30, 2011·Analytical Biochemistry·Michael D LeipoldMark Nitz

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