Classification, characterization and structural analysis of sugar nucleotidylyltransferase family of enzymes.

Sugar Nucleotidyl Transferases (SNTs) constitute a large family of enzymes that play important metabolic roles. Earlier, for one such SNT, termed N-acetylglucosamine-1-phosphate uridyltransferase- GlmU, we had established that two magnesium ions - Mg and Mg - catalyze the sugar-nucleotidyl transfer reaction. Despite a common structural framework that SNTs share, we recognized key differences around the active-site based on the analysis of available structures. Based on these differences, we had classified SNTs into two major groups, Group - I & II; and further, variation in 'Mg-stabilizing motifs' led us to sub-classify them into five distinct sub-groups. Since group specific conservation of 'Mg-stabilizing motifs' was based only for 45 available structures, here we validate this via an exhaustive analysis of 1,42,025 protein sequences. Previously, we had hypothesized that a metal-ion-catalyzed mechanism would be operative in all SNTs. Here, we validate it biochemically and establish that Mg is a strict requirement for nucleotidyl transfer reactions in every group or sub-group and that a common metal ion dependent mechanism operates in SNTs. Further, mutating Mg coordinating residue in each sub-group led to abolished catalysis, indicating an important role for both of these residues and suggest that SNTs employ variations over 'a conserved catalytic mechanism mediated by Mg ion(s)', to bring about functional diversity. This would constitute a comprehensive study to establish the catalytic mechanism across the family of SNTs.