Histidine-rich protein Hpn from Helicobacter pylori forms amyloid-like fibrils in vitro and inhibits the proliferation of gastric epithelial AGS cells

Abstract

Helicobacter pylori causes various gastric diseases, such as gastritis, peptic ulcerations, gastric cancer and mucosa-associated lymphoid tissue lymphoma. Hpn is a histidine-rich protein abundant in this bacterium and forms oligomers in physiologically relevant conditions. In this present study, Hpn oligomers were found to develop amyloid-like fibrils as confirmed by negative stain transition electron microscopy, thioflavin T and Congo red binding assays. The amyloid-like fibrils of Hpn inhibit the proliferation of gastric epithelial AGS cells through cell cycle arrest in the G2/M phase, which may be closely related to the disruption of mitochondrial bioenergetics as reflected by the significant depletion of intracellular ATP levels and the mitochondrial membrane potential. The collective data presented here shed some light on the pathologic mechanisms of H. pylori infections.

Inhibition ofHelicobacter pyloriurease activityin vivoby the synthetic nickel binding protein Hpn

“…3). This behavior confi rmed the earlier fi nding that nickel binding stabilizes the aggregated from of Hpn [11,12]. Notably, HpnA monomers were not transformed to HpnB in the presence of sodium and other cations, indicating that the metal interaction of Hpn is specifi c for nickel.…”

“…2 ). The nickel binding capacity of Hpn described earlier [10][11][12] was analyzed by co-incubation of 2 nmol Hpn with nickel agarose at increasing concentrations in phosphate buffered solution with a total volume of 30 μl. After 1 h incubation at 4 °C under shaking conditions, nickel agarose was removed by centrifugation (10 min, 4 °C, 13,000 rpm).…”

Common themes and unique proteins for the uptake and trafficking of nickel, a metal essential for the virulence of Helicobacter pylori

“…While Hpn and Hpn-2 proteins are certainly central in the nickel trafficking pathways of H. pylori and are possibly directly or indirectly involved in urease activation, their respective roles in these processes remain to be established. Recently, purified Hpn was shown to form in vitro amyloïd-like fibrils that are toxic when applied to cultured gastric epithelial cells (Ge et al, 2011). The existence and function of these fibers and the effect of nickel on their formation has yet to be demonstrated in vivo .…”

The Interaction of a Histidine‐Rich Protein Hpn with the Membrane Mimics: Implications for Pathologic Roles of Hpn in Helicobacter pylori

“…It is the first time ever that a bacterial amyloid peptide has been found to form pores in the mitochondrial membrane‐based mimics. Previously, we found that Hpn could disrupt the integrity of the mitochondrial membrane through the identification of the depletion of the intracellular ATP levels and the mitochondrial membrane potential . Our experiments involving the Laurdan and Prodan fluorescence indicated that increasing the total protein/lipid ratio leads to a less ordered and more hydrated lipid membrane structure close to the water/lipid interface of lipid bilayers modeling the mitochondrial inner membrane.…”

“…It is of interest that Aβ(1–42) is also rich in pairs of one specific amino acid, such as one pair each of His, Phe, Ile, Gly, and Val. The formation of amyloid fibrils by Hpn protein was confirmed by transition electron microscopy, and Thioflavin T and Congo red binding assays . The amyloid fibrils of Hpn inhibit the proliferation of gastric epithelial AGS cells through cell cycle arrest and disruption of mitochondrial bioenergetics .…”