biostudies-literaturePettit NNICHD NIH HHSUS National Institutes of Child Health & Human DevelopmentUS National Institutes of Genetic Medical SciencesNIGMS NIH HHS190-5https://www.ebi.ac.uk/biostudies/studies/S-EPMC3441828biostudies-literatureUnknown402(2)Escherichia coli O127:K63(B8) possesses high human blood group H (O) activity due to its O-antigen repeating unit structure. In this work, the wbiQ gene from E. coli O127:K63(B8) was expressed in E. coli BL21 (DE3) and purified as a fusion protein containing an N-terminal GST affinity tag. Using the GST-WbiQ fusion protein, the wbiQ gene was identified to encode an α1,2-fucosyltransferase using a radioactivity based assay, thin-layer chromatography assay, as well confirming product formation by using mass spectrometry and NMR spectroscopy. The fused enzyme (GST-WbiQ) has an optimal pH range from 6.5 to 7.5 and does not require the presence of a divalent metal to be enzymatically active. WbiQ displays strict substrate specificity, displaying activity only towards acceptors that contain Gal-Biochemical and biophysical research communicationsCharacterization of WbiQ: An α1,2-fucosyltransferase from Escherichia coli O127:K63(B8), and synthesis of H-type 3 blood group antigen.PMC3441828R01 HD061935R01 GM085267Wang PGPettit NMei ZHan WZhao GStyslinger TfalseCharacterization of WbiQ: An α1,2-fucosyltransferase from Escherichia coli O127:K63(B8), and synthesis of H-type 3 blood group antigen.Escherichia coli O127:K63(B8) possesses high human blood group H (O) activity due to its O-antigen repeating unit structure. In this work, the wbiQ gene from E. coli O127:K63(B8) was expressed in E. coli BL21 (DE3) and purified as a fusion protein containing an N-terminal GST affinity tag. Using the GST-WbiQ fusion protein, the wbiQ gene was identified to encode an α1,2-fucosyltransferase using a radioactivity based assay, thin-layer chromatography assay, as well confirming product formation by using mass spectrometry and NMR spectroscopy. The fused enzyme (GST-WbiQ) has an optimal pH range from 6.5 to 7.5 and does not require the presence of a divalent metal to be enzymatically active. WbiQ displays strict substrate specificity, displaying activity only towards acceptors that contain Gal-2010-01-01T00:00:00Z2010 Nov2026-05-04T20:46:06.613Z2019-03-27T00:57:56ZS-EPMC34418282080110310.1016/j.bbrc.2010.08.087