The cpsIaJ gene of Streptococcus agalactiae type la codes for beta-1,4-galactosyltransferase. In this study, the functional expression of His-tagged CpsIaJ in a baculovirus expression system was performed, because the efficient functional expression of this enzyme in Escherichia coli had been unsuccessful. Using a partially purified enzyme preparation, we found that the enzyme had a restricted substrate specificity and that the entire structure of the substrate GlcNAc beta 1-3Gal beta 1-4Glc was required for the activity. Furthermore, mutations in a conserved DXD motif caused the loss of the enzyme's activity.