Related Experiment Video
Updated: Jun 13, 2026

High-throughput Purification of Affinity-tagged Recombinant Proteins
Published on: August 26, 2012
Expression and purification of untagged and histidine-tagged folate-dependent tRNA:m5U54 methyltransferase from
Djemel Hamdane1, Stéphane Skouloubris, Hannu Myllykallio
1Laboratoire d'Enzymologie et Biochimie Structurales, CNRS, 1 Avenue de la Terrasse, 91190 Gif-sur-Yvette, France.
Abstract:
Folate-dependent tRNA m(5)U methyltransferase TrmFO is a flavoprotein that catalyzes the C(5)-methylation of uridine at position 54 in the TPsiC 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.

