The whole cell extract of Helicobacter pylori strain 26695 was treated with the hemin-agarose resin and the bound fraction was analyzed by 2-Dimensional electrophoresis. The 2-D-PAGE-displayed spots were eluted and analyzed by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS). Among the 120 spots processed, 94 protein spots were identified to represent 58 genes. Forty-five protein spots that represented thirty-four genes were newly identified in this study, including iron-containing proteins and hemin-containg proteins such as fumarate reductase, iron-sufur subunit(FrdB), ribonucleoside diphosphate reductase, beta subunit (NrdB), glutamyl-tRNA reductase (HemA), nikel-cobalt-cadnium resistance protein (NccB), and porphobilinogen deaminase (HemC).
Chromatography, Affinity* ; Electrophoresis ; Helicobacter pylori* ; Helicobacter* ; Hydroxymethylbilane Synthase ; Mass Spectrometry ; Oxidoreductases ; Proteome* ; Ribonucleoside Diphosphate Reductase ; Succinate Dehydrogenase

The nucleotide sequence of pZMO1, a small cryptic plasmid of Zymomonas mobilis ATCC10988 was determined. Analysis of 1,680 bp of sequence revealed 69% identity with Shigella sonnei plasmid, pKYM and 61% identity with Nostoc sp. ss DNA replicating plasmid. Analysis of a deduced amino acid sequence of an orf of pZMO1 revealed 75% identity and 90% similarity with the repA gene of Synechocystis sp. plasmid pCA2.4. The upstream region of the repA gene of pZMO1 possesses six directed repeat sequences and two inverted repeat sequences at downstream of the IR consensus sequence of nick region of rolling circle replication (RCR) plasmid. A typical terminator hairpin structure was found at the downstream region of repA gene. Degradation of single-stranded plasmid DNA by S1 nuclease was detected by Southern hybridization. It suggests that pZMO1 replicates by a rolling circle mechanism in Z. mobilis ATCC10988 cells.
Amino Acid Sequence ; Animals ; Base Sequence ; Consensus Sequence ; DNA ; Ecthyma, Contagious ; Inverted Repeat Sequences ; Nostoc ; Plasmids* ; Shigella sonnei ; Synechocystis ; Zymomonas*

No abstract available.
Hypertension, Renovascular*

The component of Helicobacter pylori responsible for mouse death was identified and partially characterized. Mice that were injected with H. pylori cell lysate showed pathological changes such as decreased activity, diarrhea, mild convulsion, dramatic decline of body temperature, and even death. In order to identify the lethal factor, recently isolated H. pylori strain 335 and old culture (for 10 years) of H. pylori strain 51 were used. LD50 of the cell lysate of H. pylori 335 and 51 were 338 microgram and 985 microgram, respectively, meaning the long passage of H. pylori in the laboratory might have decreased the lethal activity in the lysate. Mouse lethal activity disappeared by either treatment of cell lysate with proteinase K or heating cell lysate at 60 degrees C for 30min. Mutation analysis and genetic complementation study revealed that active urease of H. pylori is the mouse lethal factor. The recombinant H. pylori urease expressed in Escherichia coli showed si milar lethal activity.
Animals ; Body Temperature ; Complement System Proteins ; Diarrhea ; Endopeptidase K ; Escherichia coli ; Heating ; Helicobacter pylori* ; Helicobacter* ; Hot Temperature ; Lethal Dose 50 ; Mice* ; Seizures ; Urease*

As part of an initial inquiry into the function of the outer membrane proteins (OMPs) of Helicobacter pylori Korean strain 51, we have conducted an extensive proteome analysis via quadrupole time of flight (Q-TOF) mass spectrometry (MS). Fifty one OMPs of H. pylori were purified using sarcosine and resolved via two-dimensional electrophoresis with immobilized pH gradient strips. The most abundant proteins were observed in the alkaline pI regions (6.0~11.0) at molecular masses between 10~100 KDa. Here, 15 spots were identified, representing 9 types of genes (KHP0852, KHP0853, KHP1353, KHP1017, KHP0172, KHP0076, KHP0617, KHP1069, KHP0614) from the sarcosin-insoluble fraction of H. pylori 51. These may be employed in the characterization of the OMPs of H. pylori 51, which will help to identify new potential target proteins for vaccine development and drug therapy.
Electrophoresis ; Helicobacter ; Helicobacter pylori ; Mass Spectrometry ; Membrane Proteins ; Membranes ; Proteins ; Proteome ; Proton-Motive Force ; Sarcosine ; Sprains and Strains

Helicobacter pylori (H. pylori) may be one of the most common pathogen in gastrointestinal tract. Although several recent articles have reported a decline in the prevalence of H. pylori infection in both children and adults over the last several years, H. pylori infection usually occurs early in life and persists for a long time. The role of H. pylori in some digestive diseases, such as gastritis, ulcer and gastric cancer has been well established. And the possible role of H. pylori as a trigger for some extraintestinal diseases in children and adults has been considered in the last year. H. pylori infection might be associated with refractory iron deficiency anemia, idiopathic thrombocytopenic purpura, growth retardation and obesity etc, directly or indirectly. Most of the studies are classified as epidemiological, clinical researches about effects on extraintestinal manifestations after eradication, or case reports. This review describes the possibility of association with several extraintestinal diseases and H. pylori infection and their possible mechanisms based on reported studies in the world and our several studies, even though there are still many conflicting results about that.
Adult ; Anemia, Iron-Deficiency ; Child ; Gastritis ; Gastrointestinal Tract ; Helicobacter ; Helicobacter pylori ; Humans ; Obesity ; Prevalence ; Purpura, Thrombocytopenic, Idiopathic ; Stomach Neoplasms ; Ulcer

Johne's disease (JD) is a chronic, debilitating disease of ruminants including cows, and is caused by Mycobacterium avium subsp. paratuberculosis (MAP). MAP is not only important in animal husbandry, but also in public health as it is associated with the onset of Crohn's disease, a chronic inflammatory bowel disease in humans. JD, like other mycobacterial diseases including tuberculosis, is classified into different stages based on the progression of infection. In addition, development of diagnostic assays that can distinguish between subclinical and clinical stages of JD is essential to control mycobacterial infection by providing an effective treatment. For the development of novel diagnostic methods of JD, it is important to investigate and understand the mRNA expression of the various immune markers in individuals at each stage of infection. In this study, we measured the levels of Th1-type chemokines, CXCR3, CCL4, CCL5, CXCL9, CXCL10, and CXCL11 in MAP-infected bovine blood by interferon (IFN)-γ release assay (IGRA) using IFN-γ as an alternative biomarker. The association of mRNA expression patterns of these chemokines with the MAP infection stages was analyzed and IFN-γ, CCL5, and CXCL10 were found to be significantly upregulated compared to IFN-γ, the biomarker used in IGRA. Our results further indicate that IFN-γ levels significantly increased in individuals with MAP-specific antibody, and CCL5 and CXCL10 levels significantly increased in those with MAP DNA. In particular, CCL5 was significantly upregulated in individuals, in which both MAP-specific antibody and MAP DNA were detected, but the expression of CXCL10 was specifically elevated in MAP DNA-detected individuals without MAP-specific antibody.
Animal Husbandry ; Animals ; Biomarkers ; Cattle* ; Chemokines* ; Crohn Disease ; DNA ; Gene Expression* ; Humans ; Inflammatory Bowel Diseases ; Interferons ; Mycobacterium avium subsp. paratuberculosis ; Mycobacterium avium* ; Mycobacterium* ; Paratuberculosis ; Public Health ; RNA, Messenger ; Ruminants ; Transcriptome* ; Tuberculosis

BACKGROUND: The problems of coronary stent thrombosis and restenosis still remain to be solved.The glycoprotein IIb/IIIa receptor blocker, Abciximab (ReoPro), plays important roles in the treatment of high-risk patient with acute platelet-rich thrombus and in the inhibition of smooth muscle cell proliferation. The aim of this study was to determine whether the use of ReoPro-coated stents could reduce the neointimal formation in a porcine coronary stent restenosis model. METHODS: ReoPro was coated on the surface of stent by means of plasma polymerization followed by chemical grafting. Stent overdilation injury was performed with control bare stent (Group I, n=13), and ReoPro-coated stents (Group II, n=14). Follow-up quantitative coronary angiogram was performed at 4 weeks after stenting and histopathologic assessment were compared in both groups. RESULTS: The diameter stenosis by QCA between two groups was significantly higher in Group I (23+/-5 % vs. 15+/-7 %, p=0.003). On histopathologic examination, no in-stent thrombus was observed. The percent area stenosis was significantly higher in Group I than in Group II (48+/-17 % vs. 30+/-16 %, p=0.01). The area of neoinima was larger in Group I than in Group II (3.2+/-1.2 mm2 vs. 2.0+/-1.0 mm2, p=0.01). By immunocytochemistry, proliferation cell nuclear antigen indices were higher in Group I (4.2+/-2.1 %, vs 2.4+/-1.8 % p=0.03). CONCLUSION: The ReoPro-coated stent is safe and effective in the prevention of in-stent thrombus and restenosis, which may be related with the inhibition of platelet thrombus and neointimal cell proliferation.
Blood Platelets* ; Cell Proliferation ; Constriction, Pathologic ; Follow-Up Studies ; Glycoproteins* ; Humans ; Immunohistochemistry ; Myocytes, Smooth Muscle ; Neointima ; Plasma ; Polymerization ; Polymers ; Stents* ; Thrombosis ; Transplants

Helicobacter pylori, a causative agent of gastroduodenal diseases, is a Gram-negative microaerophilic bacterium. Although H. pylori locates in the microaerophilic mucous layer, the bacteria would come into contact harmful reactive oxygen species generated by host immune system. It has been reported that H. pylori harbors various defense mechanisms which can protect bacterial cells from oxygen exposure. The change of the gene expression profile of sodB-negative isogenic mutant of H. pylori 26695 was analyzed by high resolution 2-DE followed by matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and tandem MS and microarray analysis. Eighteen genes and 41 genes were upregulated and downregulated respectively, either transcriptionally or translationally. Expression levels of three genes including trxB, yxjE and ribE that were changed both on a mRNA level and on a protein level were confirmed by RT-PCR analysis. However, change of expression levels of other major antioxidants such as KatA, AhpC and NapA were not detected, which means Sod is regulated by different way from that of KatA and AhpC. Mutant study of other antioxidant proteins may give us better understanding for the regulation of stress response in H. pylori.
Antioxidants ; Bacteria ; Defense Mechanisms ; Gene Expression* ; Helicobacter pylori ; Helicobacter* ; Immune System ; Mass Spectrometry ; Microarray Analysis ; Oxygen ; Reactive Oxygen Species ; Ribes ; RNA, Messenger ; Superoxide Dismutase* ; Superoxides* ; Transcriptome

PURPOSE: The serologic diagnosis of rotaviral infections is not commonly used in clinical practice, but is used in seroepidemiologic studies. In this study, the usefulness of Escherichia coli-expressed recombinant VP6 proteins of group A rotavirus in the serodiagnosis of rotavirus infections by ELISA was evaluated. METHODS: The recombinant VP6 proteins of group A rotavirus expressed in E. coli Rosetta II strain were purified and identified. One hundred sera from 22 children (4 healthy neonates, 13 healthy children, and 5 immunocompromised children) who had serial sera samples prior to and after rotavirus infections were provided by the Gyeongsang National University Hospital, a member of the National Biobank of Korea. IgG, IgA, and IgM antibodies against rVP6 were analyzed by ELISA in all of the patients and Western blot analysis in 4 neonates. RESULTS: ELISA tests using rVP6 proteins of group A rotavirus as antigen revealed that IgG, IgA, and IgM antibodies increased after rotaviral infections in most neonates and healthy children. IgG antibodies also increased after rotaviral infections in most immunocompromised children without an adequate increase in IgM or IgA antibodies. Western blot analysis in four neonates revealed very early IgM antibody responses, even in the sera with low optical densities in ELISA tests. CONCLUSION: Our study showed that ELISA using rVP6 as an antigen is a valid diagnostic tool for seroepidemiologic studies of rotavirus infections and Western blot analysis is a sensitive test in detecting IgG, IgA, and and IgM antibodies in patients with rotavirus infections.
Antibodies ; Antibody Formation ; Blotting, Western ; Child ; Enzyme-Linked Immunosorbent Assay ; Escherichia ; Humans ; Immunoglobulin A ; Immunoglobulin G ; Immunoglobulin M ; Infant, Newborn ; Korea ; Proteins ; Rotavirus ; Rotavirus Infections ; Seroepidemiologic Studies ; Serologic Tests ; Sprains and Strains