Alkylhydroperoxide Reductase (AhpC) of Helicobacter pylori：Proteomic Analysis and Functional Characterization under Oxidative Stress
Date Issued
2006
Date
2006
Author(s)
Chuang, Ming-Hong
DOI
zh-TW
Abstract
Helicobacter pylori (Hp) is a spiral, slow growing gram-negative microaerophilic bacterium. Many studies have documented Hp is the etiological agent of gastro-duodenal diseases, such as chronic gastritis, gastric and duodenal ulcers, and gastric cancer. Furthermore, World Health Organization (WHO) has classified Hp as a class 1 carcinogen. The colonization of Hp in human stomach can induce inflammatory and immune reaction to release reactive oxygen species (ROS). Such responses appear to play a pivotal role in the survival of Hp and pathogenesis of Hp-related diseases. Therefore, this study is aimed to elucidate the influence of oxidative stress on H. pylori and characterize its underlying mechanisms and protein-expression profiles. By proteomic approaches, we have identified more than 10 proteins. Among these, AhpC and UreE are greatly reduced under long-term oxidative stress. Moreover, the sequence-homology comparison showed that the alkylhydroperoxide reductase (AhpC), an abundant antioxidant protein in Hp, is more homologous to mammalian peroxiredoxins than to eubacterial AhpC. We further demonstrated that this protein can switch its structure and function from low-molecular-weight (LMW) dimer or oligomers with peroxide reductase activity under normal microaerobic conditions or short-term (< 8 h) oxidative shock to high-molecular-weight (HMW) complexes with molecular-chaperone function after severe long-term (> 16 h) oxidative stress (20% O2) in the presence of thioredoxin (Trx).
As regards the structural and functional properties of AhpC under oxidative stress, we found that AhpC isolated from the gastric-cancerous strains possesses higher protein stability and chaperone activity than that from the duodenal-ulcerous strains under stress conditions. The expression of AhpC in different strains of H. pylori generally decreases under long-term oxidative stress as revealed by 2D-PAGE and RT-PCR. AhpC expression under short-term shock actually increases, being localized in the interior of H. pylori from strains isolated from patients of gastric cancer in contrast to its distribution on the cell surface of the microorganism from strains isolated from patients of duodenal ulcer.
Based on the significant difference between AhpC isolated from strains of gastric cancer and duodenal ulcer, we have applied ELISA and Western blotting to study the cross-reactivity of AhpC to antisera of H. pylori-infected patients with different pathological outcomes. The immunoassay revealed that antisera from tissues of patients with more extensive inflammation like that in patients of gastric ulcer or cancer react strongly with the HMW complexes of AhpC, whereas those from less extensive inflammation like patients of gastritis react weakly with the LMW oligomeric form of AhpC. Therefore it is conceivable that the antioxidant protein AhpC of H. pylori may prove to be useful as a diagnostic protein marker to monitor extents of tissue damages from patients of gastro-duodenal diseases infected by H. pylori.
Subjects
幽門桿菌
活性氧
蛋白質體學
同源性
蛋白保護子
電顯
胃癌
十二指腸潰瘍
診斷試劑
Helicobacter pylori
Proteomics
oxidative stress
2D-PAGE
RT-PCR
TEM
Western blotting
ELISA
gastric cancer
duodenal ulcer
molecular chaperone
homology
SDGs
Type
other