Bacterial UDP-N-acetylglucosamine 2-epimerase catalyzes the reversible
epimerization at C-2 of UDP-N-acetylglucosamine (UDP-GlcNAc) and thereby
provides bacteria with UDP-N-acetylmannosamine (UDP-ManNAc), the activated donor
of ManNAc residues. ManNAc is critical for several processes in bacteria,
including formation of the antiphagocytic capsular polysaccharide of pathogens
such as Streptococcus pneumoniae types 19F and 19A. We have determined the X-ray
structure (2.5 A) of UDP-GlcNAc 2-epimerase with bound UDP and identified a
previously unsuspected structural homology with the enzymes glycogen
phosphorylase and T4 phage beta-glucosyltransferase. The relationship to these
phosphoglycosyl transferases is very intriguing in terms of possible
similarities in the catalytic mechanisms. Specifically, this observation is
consistent with the proposal that the UDP-GlcNAc 2-epimerase-catalyzed
elimination and re-addition of UDP to the glycal intermediate may proceed
through a transition state with significant oxocarbenium ion-like character. The
homodimeric epimerase is composed of two similar alpha/beta/alpha sandwich
domains with the active site located in the deep cleft at the domain interface.
Comparison of the multiple copies in the asymmetric unit has revealed that the
epimerase can undergo a 10 degrees interdomain rotation that is implicated in
the regulatory mechanism. A structure-based sequence alignment has identified
several basic residues in the active site that may be involved in the proton
transfer at C-2 or stabilization of the proposed oxocarbenium ion-like
transition state. This insight into the structure of the bacterial epimerase is
applicable to the homologous N-terminal domain of the bifunctional mammalian
UDP-GlcNAc "hydrolyzing" 2-epimerase/ManNAc kinase that catalyzes the
rate-determining step in the sialic acid biosynthetic pathway.
