Sialic acid groups of protein N -glycans are important determinants of biological activity. Exposed at the end of the glycan chain, they are potential targets for glycan remodeling. Sialyltransferases (STs; EC 2.4.99) are the enzymes that catalyze the sialic acid transfer from a CMP-activated donor on to a carbohydrate acceptor in vivo. Recombinant expression of the full-length human β-galactoside α2,6 sialyltransferase I (ST6Gal-I) was hampered and therefore variants with truncated N-termini were investigated. We report on the distinct properties of two N-terminally truncated versions of ST6Gal-I, namely 89ST6Gal-I and 108ST6Gal-I, which were successfully expressed in human embryonic kidney cells. The different properties of these enzymes result most probably from the loss of interactions from helix α1 in the 108ST6Gal-I variant, which plays a role in acceptor substrate binding. The K m for N -acetyl- d -lactosamine was 10-fold increased for 108ST6Gal-I (84 mM) as compared to 89ST6Gal-I (8.3 mM). The two enzyme variants constitute a suitable tool box for the terminal modification of N -glycans. While the enzyme 89ST6Gal-I exhibited both ST (di-sialylation) and sialidase activity on a monoclonal antibody, the enzyme 108ST6Gal-I showed only ST activity with specificity for mono-sialylation.