Dataset Information

Cj1386, an atypical hemin-binding protein, mediates hemin trafficking to KatA in Campylobacter jejuni.

ABSTRACT: Catalase enzymes detoxify H2O2 by the dismutation of H2O2 into O2 and H2O through the use of hemin cofactors. While the structure and biochemical properties of catalase enzymes have been well characterized over many decades of research, it remained unclear how catalases acquire hemin. We have previously reported that Cj1386 is essential for ensuring proper hemin content in Campylobacter jejuni catalase (KatA) (A. Flint, Y. Q. Sun, and A. Stintzi, J Bacteriol 194: 334-345, 2012). In this report, an in-depth molecular characterization of Cj1386 was performed to elucidate the mechanistic details of this association. Coimmunoprecipitation assays revealed that KatA-Cj1386 transiently interact in vivo, and UV-visible spectroscopy demonstrated that purified Cj1386 protein binds hemin. Furthermore, hemin titration experiments determined that hemin binds to Cj1386 in a 1:1 ratio with hexacoordinate hemin binding. Mutagenesis of potential hemin-coordinating residues in Cj1386 showed that tyrosine 57 was essential for hemin coordination when Cj1386 was overexpressed in Escherichia coli. The importance of tyrosine 57 in hemin trafficking in vivo was confirmed by introducing the cj1386(Y57A) allele into a C. jejuni ?cj1386 mutant background. The cj1386(Y57A) mutation resulted in increased sensitivity toward H2O2 relative to the wild type, suggesting that KatA was not functional in this strain. In support of this finding, KatA immunoprecipitated from the ?cj1386+cj1386(Y57A) mutant had significantly reduced hemin content compared to that of the cj1386(WT) background. Overall, these findings indicate that Cj1386 is involved in directly trafficking hemin to KatA and that tyrosine 57 plays a key role in this function.

SUBMITTER: Flint A

PROVIDER: S-EPMC4325098 | biostudies-literature | 2015 Mar

REPOSITORIES: biostudies-literature

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Cj1386, an atypical hemin-binding protein, mediates hemin trafficking to KatA in Campylobacter jejuni.

Flint Annika A Stintzi Alain A

Journal of bacteriology 20141229 5

Catalase enzymes detoxify H2O2 by the dismutation of H2O2 into O2 and H2O through the use of hemin cofactors. While the structure and biochemical properties of catalase enzymes have been well characterized over many decades of research, it remained unclear how catalases acquire hemin. We have previously reported that Cj1386 is essential for ensuring proper hemin content in Campylobacter jejuni catalase (KatA) (A. Flint, Y. Q. Sun, and A. Stintzi, J Bacteriol 194: 334-345, 2012). In this report, ...[more]

PMID: 25548249

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Project description:BackgroundHuman Campylobacter jejuni infections are progressively rising worldwide. Information about the molecular mechanisms underlying campylobacteriosis, however, are limited. In the present study we investigated whether cytokines such as IL-23, IL-22 and IL-18, which share pivotal functions in host immunity, were involved in mediating intestinal and systemic immunopathological responses upon C. jejuni infection.Methodology/principal findingsTo assure stable infection, gnotobiotic (i.e. secondary abiotic) IL-23p19-/-, IL-22-/- and IL-18-/- mice were generated by broad-spectrum antibiotic treatment. Following peroral C. jejuni strain 81-176 infection, mice of all genotypes harbored comparably high pathogenic loads in their intestines. As compared to wildtype controls, however, IL-18-/- mice displayed less distinct C. jejuni induced sequelae as indicated by less pronounced large intestinal shrinkage and lower numbers of apoptotic cells in the colonic epithelial layer at day 8 postinfection (p.i.). Furthermore, lower colonic numbers of adaptive immune cells including regulatory T cells and B lymphocytes were accompanied by less distinct secretion of pro-inflammatory cytokines such as TNF and IFN-γ and lower IL-17A mRNA expression levels in colonic ex vivo biopsies of infected IL-18-/- as compared to wildtype mice. Upon C. jejuni infection, colonic IL-23p19 expression was up-regulated in IL-18-/- mice only, whereas IL-22 mRNA levels were lower in uninfected and infected IL-23p19-/- as well as infected IL-18-/- as compared to respective wildtype control mice. Remarkably, not only intestinal, but also systemic infection-induced immune responses were less pronounced in IL-18-/- mice as indicated by lower TNF, IFN-γ and IL-6 serum levels as compared to wildtype mice.Conclusion/significanceWe here show for the first time that IL-18 is essentially involved in mediating C. jejuni infection in the gnotobiotic mouse model. Future studies need to further unravel the underlying regulatory mechanisms orchestrating pathogen-host interaction.

Dataset Information

Cj1386, an atypical hemin-binding protein, mediates hemin trafficking to KatA in Campylobacter jejuni.

Publications

Cj1386, an atypical hemin-binding protein, mediates hemin trafficking to KatA in Campylobacter jejuni.

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