• 한국약용작물학회지
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• 제13권5호
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• pp.276-283
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• 2005

Ascorbic acid is a great antioxidant and helps protect the body against pollutants. GDP-mannose pyrophosphorylase (GMPase) is a key enzyme in manufacturing GDP-mannose, a glycosyl donor for ascorbate and cell wall biosynthesis as well as for protein glycosylation. In this study, we described molecular cloning of a full-length cDNA from Potato (Solanum tuberosum L. cv. Jasim), using tuber. The cDNA isolated encoded a GDP-mannose pyrophosphrylase. The nucleotide sequence of pGMPC showed about 95%, 89% and 80% homology with S. tuberosum (AF022716), N. tabacum (AB066279) and A. thaliana (AF076484) cDNAs clone known as GMPase, respectively. We detected the expression of GMPase using RT-PCR. The highest expression of GMPase was found in stems, and the largest amount of ascorbic acid was also presented in stems. In contrast, the leaf showed minimal level of GMPase transcript and ascorbic acid content. We propose that GMPase expression patterns were similar to the changes of ascorbic acid content in the leaves treated with diverse stresses.

• BMB Reports
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• 제42권8호
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• pp.523-528
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• 2009

Phosphomannose isomerase (PMI) catalyzes the interconversion of fructose-6-phosphate and mannose-6-phosphate in the extracellular polysaccharide (EPS) synthesis pathway. The gene encoding PMI in Sphingomonas chungbukensis DJ77 was cloned and expressed in E. coli. The pmi gene is 1,410 nucleotides long and the deduced amino acid sequence shares high homology with other bifunctional proteins that possess both PMI and GDP-mannose pyrophosphorylase (GMP) activities. The sequence analysis of PMI revealed two domains with three conserved motifs: a GMP domain at the N-terminus and a PMI domain at the C-terminus. Enzyme assays using the PMI protein confirmed its bifunctional activity. Both activities required divalent metal ions such as $Co^{2+}$, $Ca^{2+}$, $Mg^{2+}$, $Ni^{2+}$ or $Zn^{2+}$. Of these ions, $Co^{2+}$ was found to be the most effective activator of PMI. GDP-D-mannose was found to inhibit the PMI activity, suggesting feedback regulation of this pathway.

• Parasites, Hosts and Diseases
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• 제44권1호
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• pp.15-20
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• 2006

Free-living amoebae of the genus Acanthamoeba are causative agents of granulomatous amebic encephalitis and amebic keratitis. Because the virulence of Acanthamoeba culbertsoni cultured in the laboratory is restored by consecutive brain passages, we examined the genes induced in mouse brain-passaged A. culbertsoni by differential display reverse transcriptase polymerase chain reaction (DDRT-PCR). Enhanced A. culbertsoni virulence was observed during the second mouse brain passage, i.e., infected mouse mortality increased from $5\%\;to\;70\%.$ Ten cDNAs induced during mouse brain passage were identified by DDRT-PCR and this was confirmed by northern blot analysis. BlastX searches of these cDNAs indicated the upregulations of genes encoding predictive NADH-dehydrogenase, proteasomal ATPase, and GDP-mannose pyrophosphorylase B, which have previously been reported to be associated with A. culbertsoni virulence factors.

• 대한수의학회지
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• 제36권1호
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• pp.109-118
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• 1996

The Salmonella rfb gene encoding for the biosynthesis of the oligosaccharide-repeating units of the O-antigenic determinants was cloned and sequenced. A set of nucleotide primers(a forward and reverse) was selected to target a defined region of the guanosine diphospho-mannose(GDP-Man) pyrophosphorylase synthase gene : rfbM of Salmonella C serogroup. The primer set was used to develop a PCR-based rapid and specific detection system for Salmonella C1 serogroup. Amplification bands of predicted size(1,422bp) were generated from 11 different Salmonella C1 isolates. The bands were verified to be specific for the C1 serogroup by Southern blot analysis using reference homologous DNA specificity was further confirmed by the lack of reactivity with heterologous DNA derived from non-salmonella members of the family enterobacteriaeceae. A specificity of 100% was deduced along with a very high sensitivity shown by a detection limit of 1fg of a purified DNA template. The isolated DNA sequence was found to be 99.8% homologous to S montevideo but the related primers amplified with the predicted band sizes with all the Salmonella C1 serogroups tested. It is concluded that the PCR protocol based on the rfbM gene from S cholerasuis is optimal fast and specific for the detection of Salmonella C1 serogroup and also the corresponding probe is suitable for rapid detection of all Salmonella C1 serogroup DNA tested. This technology should facilitate the identification of contaminated pig products and for any other products contaminated with the Salmonalla C1 serogroup. The immediate impact of this developed method will be in the area of food safety of pig products with the potential prospect for adaptation to other food inspection technologies.