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

Abstract : Folate-dependent tRNA m5U methyltransferase TrmFO is a flavoprotein that catalyzes the C5-methylation of uridine at position 54 in the T-Psi-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.
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https://hal-polytechnique.archives-ouvertes.fr/hal-00805069
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Submitted on : Friday, July 9, 2021 - 8:21:05 PM
Last modification on : Saturday, June 25, 2022 - 10:51:59 PM
Long-term archiving on: : Sunday, October 10, 2021 - 8:00:51 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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