Growth phase-dependent composition of the Helicobacter pylori exoproteome

Abstract

Helicobacter pylori colonizes the human stomach and is associated with an increased risk of gastric cancer and peptic ulcer disease. Analysis of H. pylori protein secretion is complicated by the occurrence of bacterial autolysis. In this study, we analyzed the exoproteome of H. pylori at multiple phases of bacterial growth and identified 74 proteins that are selectively released into the extracellular space. These include proteins known to cause alterations in host cells, antigenic proteins, and additional proteins that have not yet been studied in any detail. The composition of the H. pylori exoproteome is dependent on the phase of bacterial growth. For example, the proportional abundance of the vacuolating toxin VacA in culture supernatant is higher during late growth phases than early growth phases, whereas the proportional abundance of many other proteins is higher during early growth phases. We detected marked variation in the subcellular localization of putative secreted proteins within soluble and membrane fractions derived from intact bacteria. By providing a comprehensive view of the H. pylori exoproteome, these results provide new insights into the array of secreted H. pylori proteins that may cause alterations in the gastric environment.

A Nonoligomerizing Mutant Form of Helicobacter pylori VacA Allows Structural Analysis of the p33 Domain

“…pylori is a relatively noninvasive organism that lives in the gastric mucus layer overlying gastric epithelial cells, sometimes adhering to the gastric epithelium. Many H. pylori-induced alterations in the gastric mucosa are attributable to the actions of secreted bacterial proteins on host cells (4)(5)(6). One such protein is the secreted vacuolating toxin, VacA (4,(7)(8)(9)(10).…”

“…Secretion of VacA is believed to proceed via a type V or auto-transporter mechanism (6,(34)(35)(36)(37)(38). The secreted 88-kDa VacA protein can undergo limited proteolysis to yield two domains, designated p33 and p55, which remain together after proteolysis (27,36,(39)(40)(41)(42).…”

Supporting data for analysis of the Helicobacter pylori exoproteome

“…H. pylori strain 26695 was cultured at 37 °C in room air supplemented with 5% CO 2 in a serum-free medium termed “Brucella broth filtrate”. This is a modified form of sulfite-free Brucella-cholesterol broth, prepared as described [1] . Aliquots were removed at serial time points (12, 24, 30, 36, and 48 hours post-inoculation) [1] .…”

“…Supplemental Table S5 shows an analysis of the MudPIT data in Supplemental Table S4 for 33 putative secreted proteins (identified as described in [1] ). The table shows the enrichment of individual proteins in the supernatant compared to the soluble cellular fraction, as well as membrane localization calculations at two time points.…”

“… The goal of this research was to analyze the composition of the Helicobacter pylori exoproteome at multiple phases of bacterial growth (Snider et al, 2015) [1] . H. pylori was grown in a serum-free medium and at serial time points, aliquots were centrifuged and fractionated to yield culture supernatant, a soluble cellular fraction, and a membrane fraction.…”