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Moonlighting of Helicobacter pylori catalase protects against complement-mediated killing by utilising the host molecule vitronectin.

Richter C, Mukherjee O, Ermert D, Singh B, Su YC, Agarwal V, Blom AM, Riesbeck K - Sci Rep (2016)

Bottom Line: Surprisingly, by using proteomics, we found that the hydrogen peroxide-neutralizing enzyme catalase KatA is a major Vn-binding protein.Deletion of the katA gene in three different strains resulted in impaired binding of Vn.Taken together, the virulence factor KatA is a Vn-binding protein that moonlights on the surface of H. pylori to promote bacterial evasion of host innate immunity.

View Article: PubMed Central - PubMed

Affiliation: Clinical Microbiology, Department of Translational Medicine, Lund University, SE-205 02 Malmö, Sweden.

ABSTRACT
Helicobacter pylori is an important human pathogen and a common cause of peptic ulcers and gastric cancer. Despite H. pylori provoking strong innate and adaptive immune responses, the bacterium is able to successfully establish long-term infections. Vitronectin (Vn), a component of both the extracellular matrix and plasma, is involved in many physiological processes, including regulation of the complement system. The aim of this study was to define a receptor in H. pylori that binds Vn and determine the significance of the interaction for virulence. Surprisingly, by using proteomics, we found that the hydrogen peroxide-neutralizing enzyme catalase KatA is a major Vn-binding protein. Deletion of the katA gene in three different strains resulted in impaired binding of Vn. Recombinant KatA was generated and shown to bind with high affinity to a region between heparin-binding domain 2 and 3 of Vn that differs from previously characterised bacterial binding sites on the molecule. In terms of function, KatA protected H. pylori from complement-mediated killing in a Vn-dependent manner. Taken together, the virulence factor KatA is a Vn-binding protein that moonlights on the surface of H. pylori to promote bacterial evasion of host innate immunity.

No MeSH data available.


Related in: MedlinePlus

KatA increases complement resistance in a vitronectin-dependent manner.The resistance to serum complement of KR697 wt and ΔkatA was tested in a series of assays using 5% normal human serum (NHS) of unknown anti-H. pylori antibody status (a), NHS of unknown anti-H. pylori IgG status, which was depleted from vitronectin (VDS) (b), NHS of donors negative for anti-H. pylori IgG (c), and NHS of donors positive for anti-H. pylori IgG (d). Survival rates at 15, 30 and 60 min after addition of serum were determined by counting colony forming units (CFU). Depicted is the reduction in CFU/ml in percent. Results are the mean and SE of at least three independent experiments performed in technical duplicate.
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f7: KatA increases complement resistance in a vitronectin-dependent manner.The resistance to serum complement of KR697 wt and ΔkatA was tested in a series of assays using 5% normal human serum (NHS) of unknown anti-H. pylori antibody status (a), NHS of unknown anti-H. pylori IgG status, which was depleted from vitronectin (VDS) (b), NHS of donors negative for anti-H. pylori IgG (c), and NHS of donors positive for anti-H. pylori IgG (d). Survival rates at 15, 30 and 60 min after addition of serum were determined by counting colony forming units (CFU). Depicted is the reduction in CFU/ml in percent. Results are the mean and SE of at least three independent experiments performed in technical duplicate.

Mentions: Another possible outcome of Vn binding is increased serum resistance, due to the inhibitory effect of Vn on the terminal pathway of the complement system8. Even though H. pylori does typically not enter the bloodstream, it is exposed to complement, since both complement factors and regulators, including Vn, are present in the gastric epithelium during H. pylori infection3435. Furthermore, activation of complement in the presence of H. pylori has been demonstrated in vivo and in vitro3136. We obtained normal human serum (NHS) from healthy volunteers and performed serum killing assays. The survival rates of H. pylori KR697 and the KatA-deficient KR697ΔkatA in 5% NHS were determined over a time course of 60 min (Fig. 7a). Strikingly, no killing was observed in the wild type, but rather a slight increase in colony forming units (CFU) towards the end of the assay. On the contrary, H. pylori ΔkatA showed a marked decrease of 53% already after 15 min, which was, however, not statistically significant. At 30 min CFU counts were significantly reduced to 32% when compared to the wt (p < 0.05). After 60 min, the CFU/ml was reduced to 24% of the starting value (p < 0.001). Heat inactivated control serum, did not kill any of the strains (data not shown).


Moonlighting of Helicobacter pylori catalase protects against complement-mediated killing by utilising the host molecule vitronectin.

Richter C, Mukherjee O, Ermert D, Singh B, Su YC, Agarwal V, Blom AM, Riesbeck K - Sci Rep (2016)

KatA increases complement resistance in a vitronectin-dependent manner.The resistance to serum complement of KR697 wt and ΔkatA was tested in a series of assays using 5% normal human serum (NHS) of unknown anti-H. pylori antibody status (a), NHS of unknown anti-H. pylori IgG status, which was depleted from vitronectin (VDS) (b), NHS of donors negative for anti-H. pylori IgG (c), and NHS of donors positive for anti-H. pylori IgG (d). Survival rates at 15, 30 and 60 min after addition of serum were determined by counting colony forming units (CFU). Depicted is the reduction in CFU/ml in percent. Results are the mean and SE of at least three independent experiments performed in technical duplicate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4834553&req=5

f7: KatA increases complement resistance in a vitronectin-dependent manner.The resistance to serum complement of KR697 wt and ΔkatA was tested in a series of assays using 5% normal human serum (NHS) of unknown anti-H. pylori antibody status (a), NHS of unknown anti-H. pylori IgG status, which was depleted from vitronectin (VDS) (b), NHS of donors negative for anti-H. pylori IgG (c), and NHS of donors positive for anti-H. pylori IgG (d). Survival rates at 15, 30 and 60 min after addition of serum were determined by counting colony forming units (CFU). Depicted is the reduction in CFU/ml in percent. Results are the mean and SE of at least three independent experiments performed in technical duplicate.
Mentions: Another possible outcome of Vn binding is increased serum resistance, due to the inhibitory effect of Vn on the terminal pathway of the complement system8. Even though H. pylori does typically not enter the bloodstream, it is exposed to complement, since both complement factors and regulators, including Vn, are present in the gastric epithelium during H. pylori infection3435. Furthermore, activation of complement in the presence of H. pylori has been demonstrated in vivo and in vitro3136. We obtained normal human serum (NHS) from healthy volunteers and performed serum killing assays. The survival rates of H. pylori KR697 and the KatA-deficient KR697ΔkatA in 5% NHS were determined over a time course of 60 min (Fig. 7a). Strikingly, no killing was observed in the wild type, but rather a slight increase in colony forming units (CFU) towards the end of the assay. On the contrary, H. pylori ΔkatA showed a marked decrease of 53% already after 15 min, which was, however, not statistically significant. At 30 min CFU counts were significantly reduced to 32% when compared to the wt (p < 0.05). After 60 min, the CFU/ml was reduced to 24% of the starting value (p < 0.001). Heat inactivated control serum, did not kill any of the strains (data not shown).

Bottom Line: Surprisingly, by using proteomics, we found that the hydrogen peroxide-neutralizing enzyme catalase KatA is a major Vn-binding protein.Deletion of the katA gene in three different strains resulted in impaired binding of Vn.Taken together, the virulence factor KatA is a Vn-binding protein that moonlights on the surface of H. pylori to promote bacterial evasion of host innate immunity.

View Article: PubMed Central - PubMed

Affiliation: Clinical Microbiology, Department of Translational Medicine, Lund University, SE-205 02 Malmö, Sweden.

ABSTRACT
Helicobacter pylori is an important human pathogen and a common cause of peptic ulcers and gastric cancer. Despite H. pylori provoking strong innate and adaptive immune responses, the bacterium is able to successfully establish long-term infections. Vitronectin (Vn), a component of both the extracellular matrix and plasma, is involved in many physiological processes, including regulation of the complement system. The aim of this study was to define a receptor in H. pylori that binds Vn and determine the significance of the interaction for virulence. Surprisingly, by using proteomics, we found that the hydrogen peroxide-neutralizing enzyme catalase KatA is a major Vn-binding protein. Deletion of the katA gene in three different strains resulted in impaired binding of Vn. Recombinant KatA was generated and shown to bind with high affinity to a region between heparin-binding domain 2 and 3 of Vn that differs from previously characterised bacterial binding sites on the molecule. In terms of function, KatA protected H. pylori from complement-mediated killing in a Vn-dependent manner. Taken together, the virulence factor KatA is a Vn-binding protein that moonlights on the surface of H. pylori to promote bacterial evasion of host innate immunity.

No MeSH data available.


Related in: MedlinePlus