Divergence of Biochemical Function in the HAD Superfamily: d-glycero-d-manno-Heptose-1,7-bisphosphate Phosphatase (GmhB)

d-glycero-d-manno-Heptose-1,7-bisphosphate phosphatase (GmhB) is a member of the histidinol-phosphate phosphatase (HisB) subfamily of the haloalkanoic acid dehalogenase (HAD) enzyme superfamily. GmhB supports two divergent biochemical pathways in bacteria: the d-glycero-d-manno-heptose-1α-GDP pathway (in S-layer glycoprotein biosynthesis) and the l-glycero-d-manno-heptose-1β-ADP pathway (in lipid A biosynthesis). Herein, we report the comparative analysis of substrate recognition in selected GmhB orthologs. The substrate specificity of the l-glycero-d-manno-heptose-1β-ADP pathway GmhB from Escherichia coli K-12 was evaluated using hexose and heptose bisphosphates, histidinol phosphate, and common organophosphate metabolites. Only d-glycero-d-manno-heptose 1β,7-bisphosphate (kcat/Km = 7 × 106 M−1 s−1) and d-glycero-d-manno-heptose 1α,7-bisphosphate (kcat/Km = 7 × 104 M−1 s−1) displayed physiologically significant substrate activity. 31P NMR analysis demonstrated that E. coli GmhB selectively removes the C(...