Crystal structure of the bifunctional N-acetylglucosamine 1-phosphate uridyltransferase from Escherichia coli: a paradigm for the related pyrophosphorylase superfamily

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Abstract
N‐acetylglucosamine 1‐phosphate uridyltransferase (GlmU) is a cytoplasmic bifunctional enzyme involved in the biosynthesis of the nucleotide‐activated UDP‐GlcNAc, which is an essential precursor for the biosynthetic pathways of peptidoglycan and other components in bacteria. The crystal structure of a truncated form of GlmU has been solved at 2.25 Å resolution using the multiwavelength anomalous dispersion technique and its function tested with mutagenesis studies. The molecule is composed of two distinct domains connected by a long α‐helical arm: (i) an N‐terminal domain which resembles the dinucleotide‐binding Rossmann fold; and (ii) a C‐terminal domain which adopts a left‐handed parallel β‐helix structure (LβH) as found in homologous bacterial acetyltransferases. Three GlmU molecules assemble into a trimeric arrangement with tightly packed parallel LβH domains, the long α‐helical linkers being seated on top of the arrangement and the N‐terminal domains projected away from the 3‐fold axis. In addition, the 2.3 Å resolution structure of the GlmU–UDP‐GlcNAc complex reveals the structural bases required for the uridyltransferase activity. These structures exemplify a three‐dimensional template for the development of new antibacterial agents and for studying other members of the large family of XDP‐sugar bacterial pyrophosphorylases.