Matrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF)-based cloning of enolase, ENO1, from Cryphonectria parasitica

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dc.contributor.authorM J Kim-
dc.contributor.authorH J Chung-
dc.contributor.authorS M Park-
dc.contributor.authorSung Goo Park-
dc.contributor.authorD K Chung-
dc.contributor.authorM S Yang-
dc.contributor.authorD H Kim-
dc.date.accessioned2017-04-19T09:01:17Z-
dc.date.available2017-04-19T09:01:17Z-
dc.date.issued2004-
dc.identifier.issn1017-7825-
dc.identifier.urihttps://oak.kribb.re.kr/handle/201005/6567-
dc.description.abstractOn the foundation of a database of genome sequences and protein analyses, the ability to clone a gene based on a peptide analysis is becoming more feasible and effective for identifying a specific gene and its protein product of interest. As such, the current study conducted a protein analysis using 2-D PAGE followed by MALDI-TOF and ESI-MS to identify a highly expressed gene product of C. parasitica. A distinctive and highly expressed protein spot with a molecular size of 47.2 kDa was randomly selected and MALDI-TOF MS analysis was conducted. A homology search indicated that the protein appeared to be a fungal enolase (eno1). Meanwhile, multiple alignments of fungal enolases revealed a conserved amino acid sequence, from which degenerated primers were designed. A screening of the genomic λ library of C. parasitica, using the PCR amplicon as a probe, was conducted to obtain the full-length gene, while RT-PCR was performed for the cDNA. The E. coli-expressed eno1 exhibited enolase enzymatic activity, indicating that the cloned gene encoded the C. parasitica enolase. Moreover, ESI-MS of two of the separated peptides resolved from the protein spot on 2-D PAGE revealed sequences identical to the deduced sequences, suggesting that the cloned gene indeed encoded the resolved protein spot. Northern blot analysis indicated a consistent accumulation of an eno1 transcript during the cultivation.-
dc.publisherKorea Soc-Assoc-Inst-
dc.titleMatrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF)-based cloning of enolase, ENO1, from Cryphonectria parasitica-
dc.title.alternativeMatrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF)-based cloning of enolase, ENO1, from Cryphonectria parasitica-
dc.typeArticle-
dc.citation.titleJournal of Microbiology and Biotechnology-
dc.citation.number3-
dc.citation.endPage627-
dc.citation.startPage620-
dc.citation.volume14-
dc.contributor.affiliatedAuthorSung Goo Park-
dc.contributor.alternativeName김명주-
dc.contributor.alternativeName정해종-
dc.contributor.alternativeName박승문-
dc.contributor.alternativeName박성구-
dc.contributor.alternativeName정대균-
dc.contributor.alternativeName양문식-
dc.contributor.alternativeName김대혁-
dc.identifier.bibliographicCitationJournal of Microbiology and Biotechnology, vol. 14, no. 3, pp. 620-627-
dc.subject.keyword2-D PAGE-
dc.subject.keywordC. parasitica-
dc.subject.keywordenolase-
dc.subject.keywordESI-MS-
dc.subject.keywordMALDI-TOF-
dc.subject.local2-D PAGE-
dc.subject.local2D PAGE-
dc.subject.localC. parasitica-
dc.subject.localenolase-
dc.subject.localESI-MS-
dc.subject.localMALDI-TOF-
dc.subject.localMALDITOF-
dc.description.journalClassY-
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Division of Biomedical Research > Disease Target Structure Research Center > 1. Journal Articles
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