Cloning, expression, and purification of the general stress protein YhbO from Escherichia coli

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dc.contributor.author Abdallah, Jad
dc.contributor.author Kern, Renee
dc.contributor.author Malki, Abderrahim
dc.contributor.author Eckey, Viola
dc.contributor.author Richarme, Gilbert
dc.date.accessioned 2016-07-19T09:26:50Z
dc.date.available 2016-07-19T09:26:50Z
dc.date.copyright 2006 en_US
dc.date.issued 2016-07-19
dc.identifier.issn 1046-5928 en_US
dc.identifier.uri http://hdl.handle.net/10725/4165
dc.description.abstract We cloned, expressed, and purified the Escherichia coli yhbO gene product, which is an aminoacid sequence homolog to the Bacillus subtilis general stress protein 18 (the yfkM gene product), the Pyrococcus furiosus intracellular protease PfpI, and the human Parkinson disease protein DJ-1. The gene coding for YhbO was generated by amplifying the yhbO gene from E. coli by polymerase chain reaction. It was inserted into the expression plasmid pET-21a, under the transcriptional control of the bacteriophage T7 promoter and lac operator. A BL21 (DE3) E. coli strain transformed with the YhbO-expression vector, pET-21a-yhbO, accumulates large amounts of a soluble protein with a molecular mass of 20 kDa in SDS–PAGE that matches the expected YhbO molecular weight. YhbO was purified to homogeneity by ion exchange chromatography and hydroxyapatite chromatography, and its identity was confirmed by N-terminal sequencing and mass spectrometry analysis. The native protein exists in monomeric, trimeric, and hexameric forms. We also report a strong sequence homology between YhbO and the general stress protein YfkM (64% identities), which suggests that YhbO is a stress protein, and a strong structural homology between YhbO and the Pyrococcus horikoshii intracellular protease PhpI. We could not, however, detect any proteolytic or peptidolytic activity of YhbO, using classical biochemical substrates. en_US
dc.language.iso en en_US
dc.title Cloning, expression, and purification of the general stress protein YhbO from Escherichia coli en_US
dc.type Article en_US
dc.description.version Published en_US
dc.author.school SOP en_US
dc.author.idnumber 200703820 en_US
dc.author.department N/A en_US
dc.author.department Pharmaceutical Sciences Department en_US
dc.description.embargo N/A en_US
dc.relation.journal Protein Expression and Purification en_US
dc.journal.volume 47 en_US
dc.journal.issue 2 en_US
dc.article.pages 455-460 en_US
dc.keywords Escherichia coli en_US
dc.keywords General stress protein YhbO en_US
dc.keywords ThiJ superfamily en_US
dc.keywords DJ-1 en_US
dc.identifier.doi http://dx.doi.org/10.1016/j.pep.2005.11.011 en_US
dc.identifier.ctation Abdallah, J., Kern, R., Malki, A., Eckey, V., & Richarme, G. (2006). Cloning, expression, and purification of the general stress protein YhbO from Escherichia coli. Protein expression and purification, 47(2), 455-460. en_US
dc.author.email jabdallah@lau.edu.lb
dc.identifier.tou http://libraries.lau.edu.lb/research/laur/terms-of-use/articles.php en_US
dc.identifier.url http://www.sciencedirect.com/science/article/pii/S1046592805004018 en_US
dc.orcid.id https://orcid.org/0000-0001-5267-4953 en_US

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