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Expression and purification of untagged and histidine-tagged folate-dependent tRNA:m5U54 methyltransferase from Bacillus subtilis

Djemel Hamdane 1 Stéphane Skouloubris 2 Hannu Myllykallio 3 Béatrice Golinelli-Pimpaneau 1
1 LEBS - Laboratoire d'Enzymologie et Biochimie Structurales
2 IGM - Institut de génétique et microbiologie [Orsay]
3 LOB - Laboratoire d'optique et biosciences
Abstract : Folate-dependent tRNA m5U methyltransferase TrmFO is a flavoprotein that catalyzes the C5-methylation of uridine at position 54 in the T?C loop of tRNA in several bacteria. Here we report the cloning and optimization of expression in Escherichia coli BL21 (DE3) of untagged, N-terminus, C-terminus (His)6-tagged TrmFO from Bacillus subtilis. Tagged and untagged TrmFO were purified to homogeneity by metal affinity or ion exchange and heparin affinity, respectively, followed by size-exclusion chromatography. The tag did not significantly alter the expression level, flavin content, activity and secondary structure of the protein. Cop. 2010 Elsevier Inc. All rights reserved.
Keywords : Expression Purification Recombinant TrmFO protein Flavoenzyme RNA methyltransferase 5-Methyluridine Histidine tag
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Journal articles
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https://hal-polytechnique.archives-ouvertes.fr/hal-00805069
Contributor : Denis Roura <>
Submitted on : Thursday, April 4, 2013 - 10:55:28 AM
Last modification on : Thursday, November 19, 2020 - 1:34:04 PM

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Djemel Hamdane, Stéphane Skouloubris, Hannu Myllykallio, Béatrice Golinelli-Pimpaneau. Expression and purification of untagged and histidine-tagged folate-dependent tRNA:m5U54 methyltransferase from Bacillus subtilis. Protein Expression and Purification, Elsevier, 2010, 73 (1), pp.83-89. ⟨10.1016/j.pep.2010.04.013⟩. ⟨hal-00805069⟩

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