Fig. 4: In vitro biochemical analyses of enzymes OngA, OngB, OngC and KdgK.

a Negative-mode Q-TOF-MS spectrum of products generated by the recombinant OngA acting on GlcNAc-GlcNAc1A. Q-TOF-MS spectrum of GlcNAc-GlcNAc1A only as the control was shown in Supplementary Fig. 12a. DP2_ox, GlcNAc-GlcNAc1A. b Negative-mode Q-TOF-MS spectrum of products generated by the recombinant OngB acting on GlcNAc1A. Q-TOF-MS spectrum of GlcNAc1A only as the control was shown in Supplementary Fig. 12b. c Negative-mode Q-TOF-MS spectrum of products generated by the recombinant enzymes, OngB and OngC, acting on GlcNAc1A. Q-TOF-MS spectrum of KDG only as the control was shown in Supplementary Fig. 12c. d Negative-mode Q-TOF-MS spectrum of products generated by OngB, OngC and KdgK successively acting on GlcNAc1A. Q-TOF-MS spectrum of KDG-6-P only as the control was shown in Supplementary Fig. 12d. In a-d all labeled MS peaks refer to [M-H]^- ions unless noted otherwise (e.g. [GlcNAc+Cl]^- ion). Theoretical masses of relevant products are listed in Supplementary Table 4. In a-d, the graphs show a representative MS spectrum of at least three independent replicates. e Maximum-likelihood tree of OngB and its homologs and other de-N-acetylases. All homologs to OngB (colored in red) are from the ong clusters of their respective bacterial strains. CE carbohydrate esterase, DA deacetylase, PGN peptidoglycan, Acetyl-D-Glu, acetyl-D-glutamate. f Neighbor-joining tree of OngC and its homologs and characterized pyridoxal 5-phosphate (PLP)-dependent enzymes. All homologs to OngC (colored in red) are from the ong clusters of their respective bacterial strains. Source data are provided as a Source Data file.