Journal List > J Bacteriol Virol > v.37(4) > 1033880

Lee, Kim, Park, Kwon, Song, Bae, Youn, Seo, Baik, Lee, Cho, Rhee, and Kang: Comparison of Proteome Component of Helicobacter pylori in Different Atmospheric CO2 Concentration

Abstract

Helicobacter pylori is a spiral, slow growing gram-negative microaerophilic bacterium. It has been shown to be the etiological agent of gastroduodenal diseases, such as chronic gastritis, gastric and duodenal ulcers, and gastric cancer. General culture condition of H. pylori is 5% O2, 10% CO2 and 100% humid atmosphere. We have compared proliferation protein expression profile of H. pylori incubated under normal microaerophilic (10% CO2) and environment stress (4% CO2, 18% CO2) conditions. H. pylori cultured under environment stress displayed coccoid morphology and time-dependent decrease in proliferation. We have further compared the protein expression profiles of H. pylori under normal growing and environment stress conditions by a global proteomic analysis, which includes high-resolution 2-DE followed by matrix-assisted laser desorption/ionization time of flight and nanoelectrospray/tandem mass spectrometry. In total, 42 protein spots were found to be up- or down-regulated by more than 2-fold under environment stress conditions. Of the 42 protein spots processed, 27 spots were identified; they represented 19 genes, including 2 kinds of hypothetical proteins.

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Figure 1.
Morphology change of H. pylori. Original culture grown under 5% O2 and 10% CO2 (A) were transferred to the new medium and cultured 3 more days under 4% CO2 (B), 10% CO2 (C) and 18% CO2 (D).
jbv-37-213f1.tif
Figure 2.
Two-DE of whole cell protein extracts (200 μg) of H. pylori. The proteins were separated on an IPG strip of pH 5.0∼8.0 and subsequently on a 12.5% SDS-PAGE gel and then detected by silver staining. The original gel size is 17×20×0.12 cm. The numbers on the left indicate molecular mass markers and pI values at the top of the panel. (A) 10% CO2, 5% O2 (B) 5% CO2, 5% O2 (C) 18% CO2, 5% O2
jbv-37-213f2.tif
Figure 3.
Comparison of expression intensity in over two fold increased or decreased protein spots in CO2 4% and/or 18%. X axis and Y axis represent spot number and spot intensity, respectively. SSP No is standard spot number in PDQUEST program.
jbv-37-213f3.tif
Table 1.
Identified up or down-regulated proteins under various CO2 concentration of H. pylori strain 26695 by using tandem ms/ms
SSP No Proteins name Accession no MW pI Amino acid seq Score HP No
0506e Translation elongation factor EF-Tu (TufB) gi:2314366 43647.7 4.93 R.ELLSAYEFPGDDTPIVAGSALR.A 66 1205
          K.TFLMPVEDVFSIAGR.G 19  
1011c 3-dehydroquinase type II (AroQ) gi:2314182 18483.25 4.93 K.ILVIQGPNLNMLGHR.D 65 1038
          K.QGNLDVELEFFQTNFEGEIIDK.I 72  
          R.GLSLAGNQVLTR.T 63  
2104b Conserved hypothetical protein gi:2314773 28417.54 5.24 R.QALSAATLTLFK.M 46 1588
          K.TETTLIEQNMLSK.I 73  
          R.AQDFEILTSR.L 32  
2901a Polynucleotide phosphorylase (Pnp) gi:2314375 76894.54 5.34 K.NQYFDEIK.G 45 1213
          K.ESLNMIEMR.S 41  
3204b Thioredoxin reductase (TrxB) gi:2313959 33538.22 6.25 R.ELVVPGFFIFVGYDVNNAVLK.Q 53 0825
          R.LVQVVDIEK.M 48  
4205b Chemotaxis protein (CheV) gi:2313092 36587.05 6.19 K.DLYAVNVFK.I 19 0019
          R.ELTIPLIDMK.K 23  
5801b Threonyl-tRNA synthetase (ThrS) gi:2313207 70239.64 6.36 R.IYGIAFATK.E 30 0123
          K.LAGAYLGGDENNEMLI.R 74  
5901a ATP-dependent protease binding subunit (ClpB) gi:2314182 96683.36 6.27 K.ALAQFLFDSDKNLIR.I 27 0264
5911c ATP-dependent protease binding subunit (ClpB) gi:2314182 96683.36 6.27 K.ALAQFLFDSDKNLIR.I 25 0264
6302b ATP-binding protein (Mpr) gi:2313292 45197.63 8.58 K.VGLLDADVYGPNIPR.M 74 0207
6801e Hydantoin utilization protein A (HyuA) gi:2313818 78532.79 6.94 K.LLPGNEVIGPAIVESDATTFVIPK.G 33 0695
7001d Modulator of drug activity (Mda66) gi:2313748 21604.61 7.16 K.NPQVEQYLNSLTTHLR.Q 24 0630
          R.TIVSGPIGGVIGSK.L 39  
7702e Hydantoin utilization protein A (HyuA) gi:2313818 78532.79 6.94 R.LVLSLPLVAMDSVGAGAGSFVR.I 22 0695
          K.IIQDAWDELTLK.V 50  
          R.GVVATQKPVIPVEK.E 24  
7705e Hydantoin utilization protein A (HyuA) gi:2313818 78532.79 6.94 R.TIVSGPIGGVIGSK.L 59 0695
          K.YDDPLIPLKR.I 44  
          R.TIVSGPIGGVIGSK.L 31  
7801e Hydantoin utilization protein A (HyuA) gi:2313818 78532.79 6.94 R.LVLSLPLVAMDSVGAGAGSFVR.I 26 0695
          K.IIQDAWDELTLK.V 24  
8102b Uease alpha subunit (UreA) gi:2313154 26539.49 8.96 R.IFGFNALVDR.Q 71 0073
          K.LNYVEAVALISAHIMEEAR.A 18  
          K.LLAPILPLVK.E 26  
8603f Conserved hypothetical iron-sulfur protein gi:2313222 54236.31 7.04 K.DGDEANEIIYNLAK.E 53 0138
          R.LDEYLELFEK.N 48,  
8702a,e Hydantoin utilization protein A (HyuA) gi:2313818 78532.79 6.94 R.GVVATQKPVIPVEK.E 25, −0695
          R.TIVSGPIGGVIGSK.L 68,  
               
8801a Glucose inhibited divistion protein (GidA) gi:2313303 69683.72 7.13 R.LGLMEEDFYK.E 38, −0213
          K.GIPGLSLEAVEK.L 42,  
8903a,e N-methylhydantoinase gi:2313819 86511.66 6.95 R.TVLLVDEK. 31, −0696
          K.GGPGFGDPIER.D 45,  

a) Increased protein spots in 4%-CO2.

b) Increased protein spots in 18%-CO2.

c) Increased protein spots in both 4% and 18%-CO2.

d) Decreased protein spots in 4%-CO2.

e) Decreased protein spots in 18%-CO2.

f) Decreased protein spots in both 4% and 18%-CO2

Table 2.
Identified up or down-regulated proteins under various CO2 concentration of H. pylori strain 26695 by using MALDI-TOF MS
SSP No Proteins name Accession no MW pI HP no
5001b Co-chaperone (GroES) gi:2313085 12990.87 6.59 0011
2906a NADH-ubiquinone oxidoreductase, NQO3 subunit (NQO3) gi:2314431 94229.31 5.12 1266
3903a Protein translocase subunit (SecA) gi:2313910 99083.31 5.65 0786
3909a Protein translocase subunit (SecA) gi:2313910 99083.31 5.65 0786
4107c Alkyl hydroperoxide reductase (AhpC) gi:2314747 22235.57 6.25 1563

a) Increased protein spots in 4%-CO2.

b) Increased protein spots in 18%-CO2.

c) Decreased protein spots in 4%-CO2

Table 3.
Identified proteins classification by functional annotation of using COGs
Proteins name GOG No Functional annotation Code
3-dehydroquinase type II (AroQ) 0757 E
Modulator of drug activity (Mda66) 2249 R
Polynucleotide phosphorylase (Pnp) 1185 J
NADH-ubiquinone oxidoreductase, NQO3 subunit (NQO3) 1034 C
Uease alpha subunit (UureA) 0831 E
protein translocase subunit (SecA) 0653 N
ATP-dependent protease binding subunit (ClpB) 0542 O
Thioredoxin reductase (TrxB) 0492 O
ATP-binding protein (Mpr) 0489 D
Alkyl hydroperoxide reductase (AhpC) 0450 O
Glucose inhibited divistion protein (GidA) 0445 D
Threonyl-tRNA synthetase (ThrS) 0441 J
Co-chaperone (GroES) 0234 O
N-methylhydantoinase 0146 E
Hydantoin utilization protein A (HyuA) 0145 E
Translation elongation factor EF-Tu (TufB) 0050 J, E

C: Energy production and conversion

D: Cell division and chromosome partitioning

E: Amino acid transport and metabolism

J: Translation, ribosomal structure and biogenesis

N: Cell motility and secretion

O: Posttranslational modification, protein turnover, chaperones

R: General function prediction only

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