Product Name Recombinant Human CMP-N-acetylneuraminate beta-galactosamide-alpha-2,6-sialyltransferase (ST6GAL1)

Catalog Number #0002Alternate Names beta-galactosamide alpha-2,6-sialyltransferase; ST6Gal I; alpha 2,6-ST 1; B-cell antigen

CD75; ST6 beta-galactosamide alpha-2,6-sialyltranferase 1; ST6 N-acetylgalactosaminide alpha-2,6-sialyltransferase 1

Substrate Specificity Human Beta-galactoside alpha-2,6-sialyltransferase 1 (ST6Gal I) forms NeuAc 𝝰2,6-Gal linkages in N-linked glycans. ST6GAL1 adds an 𝝰2,6-linked sialic acid residue to Gal𝝱1,4-GlcNAc structures found on the N-linked glycan chains of glycoproteins [1].

References References: [1] Kitazume, S. (2013) "Chapter 63: ST6 Beta-Galacotside Alpha-2,6-Sialyltransferase 1 (ST6GAL1)" in Handbook of Glycosyltransferases and Related Genes, 2nd edition.

Expression Host HEK293Species of expressed protein HumanGene ID 6480Protein RefSeq NP_775323Uniprot P15907Region Expressed AA 75-406Expressed Protein Sequence RQTLGSLRGLAKAKPEASFQVWNKDSSSKNLIPRLQKIWKNYLSMNKYKVSYKGPGPGIKFS

AEALRCHLRDHVNVSMVEVTDFPFNTSEWEGYLPKESIRTKAGPWGRCAVVSSAGSLKSSQLGREIDDHDAVLRFNGAPTANFQQDVGTKTTIRLMNSQLVTTEKRFLKDSLYNEGILIVWDPSVYHSDIPKWYQNPDYNFFNNYKTYRKLHPNQPFYILKPQMPWELWDILQEISPEEIQPNPPSSGMLGIIIMMTLCDQVDIYEFLPSKRKTDVCYYYQKFFDSACTMGAYHPLLYEKNLVKHLNQGTDEDIYLLGKATLPGFRTIHC

Tag(s) N-terminal 6xHis, GFPSpecific Activity Specific Activity is ≥0.5 µmol/min/mg, as measured under the conditions described below.Purity (%) >95%, by SDS_PAGE under reducing conditions and visualized by Coomassie Blue stain.Formulation Supplied as a 0.2𝞵m filtered soulution in 20mM HEPES and 100mM NaCl buffer, pH 7.0,

with 10% Glycerol and 0.05 % NaN3 as preservative.

Concentration 1 µg/µlSDS-Page Size ~60-70kDaSDS-PAGE image

250150100

75

50

37

2515

10

Activity Measured by the ability to transfer the sugar from CMP-Neu5Ac and generate CMPAssay Buffer 100mM MES, pH 6.5

Donor Substrate CMP-Neu5Ac (0.25 mM, Nacalai Tesque Inc.) Acceptor Substate N-acetylactosamine (2.4 mM, Sigma)Coupling Enzyme 5-nucleotidase (CD73) Detection Kit Malachite Green Phosphate Detection Kit (R&D Systems, Minneapolis, MN)Assay Steps

1) Prepare a 50µl reaction volume containing: 100mM MES (pH 6.5), 0.1µg of recombinant ST6GAL1, CD73 (0.5 ng/µl), N -acetyllactosamine (2.4 mM), and CMP-Neu5Ac (0.25 mM) in a microfuge tube.

2) Incubate at 37C° for 30 min.3) Add 10 µl of malachite green phosphate detection reagent A and incubate for 10 min at 25

°C, then add 10 µl of reagent B and incubate for 20 min at 25 °C

4) Read absorbance using a Spectra MAX 190 spectrophotometer (Molecular Devices) at 620 nm

Std Curve Follow protocol for generating a Phosphate standard curve using the phosphate standard contained in the Malachite Green Phosphate Detection Kit (R&D Systems)

Specific Actifity calc Specific Activity (pmol/min/ug)= [Phosphate released*(nmol) x (1000 pmol/nmol)] / [Incubation time (min) x amount of enzyme (ug)] Specific Activity was calculated using the standard curve plotted in GraphPad Prism 6 (GraphPad Software)

Shipping conditions This product is shipped as 0.2µm filtered product on dry ice. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage cond6 months 6 months if stored at -80C. Avoid repeated freeze thaws.