Growth‐induced changes in the proteome of Helicobacter pylori

Abstract

Helicobacter pylori is a major human pathogen that is responsible for a number of gastrointestinal infections. We have used 2-DE to characterise protein synthesis in bacteria grown either on solid agar-based media or in each of two broth culture media (Brucella and brain heart infusion (BHI) broth). Significant differences were observed in the proteomes of bacteria grown either on agar-based or in broth media. Major changes in protein abundance were identified using principal component analysis (PCA), which delineated the profiles derived for the three key growth conditions (i.e. agar plates, Brucella and BHI broth). Proteins detected across the gel series were identified by peptide mass mapping and Edman sequencing. A number of proteins associated with protein synthesis in general as well as specific amino acid synthesis were depressed in broth-grown bacteria compared to plate-grown bacteria. A similar reduction was also observed in the abundance of proteins involved in detoxification. Two of the most abundant spots, identified as UreB and GroEL, in plate-grown bacteria showed a >140-fold drop in abundance in bacteria grown in Brucella broth compared to bacteria grown on agar plates. Two protein spots induced in bacteria grown in broth culture were both identified as glyceraldehyde 3-phosphate dehydrogenase based on their N-terminal amino acid sequences derived by Edman degradation. The underlying causes of the changes in the proteins abundance were not clear, but it was likely that a significant proportion of the changes were due to the alkaline pH of the broth culture media.

Ovine corpus luteum proteins, with functions including oxidative stress and lipid metabolism, show complex alterations during implantation

“…Equal amounts of protein from each ewe, in each group, were combined to make five protein pools as follows: cyclic ewes on days 12 (C12) and 16 (C16) and pregnant ewes on days 12 (P12), 16 (P16), and 20 (P20). 2DE was performed as described previously (Cash & Kroll 2003, Uwins et al 2006. For first dimension separation, 200 mg total protein from CL lysate was loaded on 11 cm, immobilised, non-linear pH gradient strips of pH 3-10 (Bio-Rad).…”

“…In order to identify proteins, spots were excised from stained gels and subjected to in-gel trypsin digestion as described previously (Uwins et al 2006). The peptide fragment mass spectra were acquired on a PerSeptive Biosystems Voyager-DE STR MALDI-TOF mass spectrometer operated in the reflection delayed extraction mode.…”

Proteomic Analysis of Helicobacter pylori Whole Cell Proteins using the Narrow Range IPG Strips

“…Of them, 64 proteins like urease beta subunit, 60 kDa chaperonin, and alkyl hydroperoxide reductase had been identified in the previous studies (4,10,18,20,21,23,29,33).…”

“…The genome of H. pylori has been determined to contain about 1,590 open reading frames (ORFs) (1,26,32). Up to date, more than 250 have been identified to list as proteome components of H. pylori (4,10,18,20,21,23,29,33), revealing more intensive exploration needed to construct proteomic information of H. pylori. Actually, most spots appeared so crowded in the region of pI 4.5~8.0 that they were inseparable in 2-DE using the broad range IPG strip.…”

Stable isotope labeling by amino acids in cell culture based proteomics reveals differences in protein abundances between spiral and coccoid forms of the gastric pathogen Helicobacter pylori

“…These studies investigated altered protein abundances in response to oxidative stress [9,23], mice colonization [24], gastric epithelial cell apoptosis [25], growth conditions [26], acidic stress [27] and iron uptake [28,29]. However, all of these proteomic studies were performed by two-dimensional gel electrophoresis (2D-PAGE) and therefore could only cover up to 37% of the H. pylori proteome [23][24][25][26][27][28][29][30][31][32].…”