Figure 4. The activity of BT1044^GH18 on immunoglobulin substrates and structural of the enzyme.

Activity of BT1044^GH18 against a, human serum IgG b, human serum IgA and c, human colostrum IgA is shown in the presence of BT0455^GH33 sialidase. The products with a β1,4 bisecting GlcNAc are indicated to by a white asterisk. The results are representative of at least three independent replicates. d, Cartoon depiction of the predicted orientation of BT1044^GH18 on the cell surface of Bt showing the attachment to the cell surface through a lipid anchor. Loops 1-7 on the surface of the enzyme with the active site are coloured yellow, green, orange, blue, magenta, purple and dark green, respectively. e, The surface of the BT1044^GH18 crystal structure is shown again with the same loop colouring and the two catalytic residues are in red. For comparison, the previously published crystal structure of the CNG-active enzyme EndoF3 (1EOM) from Elizabethkingia meningoseptica is shown (cyan). The conserved residues from the DxxDxDxE motif have been coloured orange and the approximate +1 (GlcNAc) and +1’ (fucose) subsites are indicated to by white and red dashed lines, respectively. BT1044^GH18 has more of a groove (white dashed box) in the equivalent section. The area where a bisecting GlcNAc would be predicted to sit is indicated to by an asterisk. This space is occupied in Endo F3, whereas in BT1044^GH18 there is more space to accommodate this sugar, reflecting their respective activities (see Supplementary Fig. 21 for more details). The results are representative of at least three independent replicates.