Abstract
BackgroundUDP-GlcNAc 2-epimerase/ManNAc 6-kinase, GNE, is a bi-functional enzyme that plays a key role in sialic acid biosynthesis. Mutations of the GNE protein cause sialurea or autosomal recessive inclusion body myopathy/Nonaka myopathy. GNE is the only human protein that contains a kinase domain belonging to the ROK (repressor, ORF, kinase) family.Principal FindingsWe solved the structure of the GNE kinase domain in the ligand-free state. The protein exists predominantly as a dimer in solution, with small populations of monomer and higher-order oligomer in equilibrium with the dimer. Crystal packing analysis reveals the existence of a crystallographic hexamer, and that the kinase domain dimerizes through the C-lobe subdomain. Mapping of disease-related missense mutations onto the kinase domain structure revealed that the mutation sites could be classified into four different groups based on the location – dimer interface, interlobar helices, protein surface, or within other secondary structural elements.ConclusionsThe crystal structure of the kinase domain of GNE provides a structural basis for understanding disease-causing mutations and a model of hexameric wild type full length enzyme.Enhanced Version This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.
Highlights
Sialic acids are N- or O- substituted terminal monosaccharides with a nine-carbon backbone highly expressed on eukaryotic cell surfaces [1]
The crystal structure of the kinase domain of GlcNAc 2-epimerase/N-acetylmannosamine kinase (GNE) provides a structural basis for understanding diseasecausing mutations and a model of hexameric wild type full length enzyme
Neu5Ac is synthesized in the cytosol from UDP-N-acetylglucosamine (UDP-GlcNAc) by four consecutive reactions; and UDP-GlcNAc is a derivative of fructose-6phosphate and the end-product of the hexosamine biosynthesis pathway (Figure 1)
Summary
Sialic acids are N- or O- substituted terminal monosaccharides with a nine-carbon backbone highly expressed on eukaryotic cell surfaces [1]. The first two steps of the biosynthesis of Neu5Ac from UDPGlcNAc are catalyzed by the bi-functional enzyme UDP-GlcNAc 2-epimerase/N-acetylmannosamine kinase (GNE). The epimerase domain converts UDP-GlcNAc to Nacetylmannosamine (ManNAc), which is phosphorylated at the 6 position by the kinase domain. The kinase domain belongs to the ROK (Repressor, ORF, Kinase) family. The ROK family consists of a set of bacterial proteins that include repressors for sugar catabolic operons, and sugar kinases [8]. Gne is the only known gene in the entire human genome that encodes a ROK domain-containing protein. UDP-GlcNAc 2-epimerase/ManNAc 6-kinase, GNE, is a bi-functional enzyme that plays a key role in sialic acid biosynthesis. GNE is the only human protein that contains a kinase domain belonging to the ROK (repressor, ORF, kinase) family
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