Crystallization (Comment) | Organism |
---|---|
enzyme in complex with UDP-GlcNAc, X-ray diffraction crystal structure determination and analysis at 2.7 A resolution. Only UDP is observed in the active site of each monomer despite the use of UDP-GlcNAc in crystallization, presumably the substrate has been converted to ManNAc and UDP before it is trapped in the crystal | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
D112A | site-directed mutagenesis, the mutant shows 97.7% reduced activity compared to the wild-type enzyme | Homo sapiens |
D143A | site-directed mutagenesis, inactive mutant | Homo sapiens |
E134A | site-directed mutagenesis, inactive mutant | Homo sapiens |
R113A | site-directed mutagenesis, inactive mutant | Homo sapiens |
S302A | site-directed mutagenesis, the mutant shows 87.1% reduced activity compared to the wild-type enzyme | Homo sapiens |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
CMP-Neu5Ac | feedback inhibition, cooperative binding mode and binding structure, overview | Homo sapiens | |
UDP | the binding mode of UDP reveals unique interactions with the hydrolyzing epimerase, binding structure, overview. The base is sandwiched between the side chains of Arg19 and Phe287, while making two hydrogen bonds to the backbone of Val282. The two hydroxyl groups of ribose are hydrogen bonded to the side chains of Ser23 and Glu307. The diphosphate forms salt bridges to Arg19, Arg113 and Arg321. It also interacts with His220 and Asn253 through hydrogen bonds, as well as the two consecutive Ser301 and Ser302 at the N-terminus of helix alpha12 | Homo sapiens |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0005 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant mutant D112A, pH 7.5, 37°C | Homo sapiens | |
0.0121 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant mutant S302A, pH 7.5, 37°C | Homo sapiens | |
0.0331 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant wild-type enzyme, pH 7.5, 37°C | Homo sapiens |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
UDP-N-acetyl-alpha-D-glucosamine + H2O | Homo sapiens | - |
N-acetyl-D-mannosamine + UDP | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | Q9Y223 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
UDP-N-acetyl-alpha-D-glucosamine + H2O | - |
Homo sapiens | N-acetyl-D-mannosamine + UDP | - |
? |
Subunits | Comment | Organism |
---|---|---|
dimer | - |
Homo sapiens |
More | the full-length enzyme can form a dimer or tetramer, depending on the presence of UDP-GlcNAc and CMP-Neu5Ac. GNE forms a closed tetramer in the presence of UDP-GlcNAc and CMP-Neu5Ac. The complex crystal structure of the N-terminal epimerase part of human GNE shows a tetramer in which UDP binds to the active site and CMP-Neu5Ac binds to the dimer-dimer interface. The enzyme is locked in a tightly closed conformation. Each dimer further forms a tetramer with a crystallographic dyad-related dimer | Homo sapiens |
tetramer | dimer of dimers | Homo sapiens |
Synonyms | Comment | Organism |
---|---|---|
GNE | - |
Homo sapiens |
UDP-GlcNAc 2-epimerase | - |
Homo sapiens |
UDP-GlcNAc 2-epimerase/ManNAc kinase | - |
Homo sapiens |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
37 | - |
assay at | Homo sapiens |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.076 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant mutant D112A, pH 7.5, 37°C | Homo sapiens | |
0.791 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant mutant S302A, pH 7.5, 37°C | Homo sapiens | |
11.8 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant wild-type enzyme, pH 7.5, 37°C | Homo sapiens |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Homo sapiens |
General Information | Comment | Organism |
---|---|---|
malfunction | defective GNE inhibition by CMP-Neu5Ac causes cytoplasmic accumulation and increased excretion of free sialic acid. Sialuria is an autosomal dominant disorder which is related to GNE mutation in one of the two arginine residues 263 and 266 (R263L, R266Q or R266W), the mutations in Arg263 and Arg266 can cause sialuria by hindering the enzyme inhibition through CMP-Neu5Ac binding | Homo sapiens |
metabolism | sialic acid biosynthesis in mammals starts by converting UDP-GluNAc into UDP and ManNAc, followed by phosphorylation of ManNAc at the sixth position, catalysis of both reactions is carried out by the bifunctional enzyme GNE. Regulation of cell-surface sialyation level by binding to the downstream product CMP-Neu5Ac. The feedback inhibition is highly positively cooperative and it does not affect the ManNAc kinase activity | Homo sapiens |
additional information | the comparison of the UDP-binding modes of the non-hydrolyzing and hydrolyzing UDP-GlcNAc epimerases might explain the mechanistic difference. While the epimerization reactions of both enzymes are similar, Arg113 and Ser302 of GNE are likely involved in product hydrolysis. Full-length modelling suggests a channel for ManNAc trafficking within the bifunctional enzyme, possible epimerase-kinase channel | Homo sapiens |
physiological function | the bifunctional enzyme UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE) plays a key role in sialic acid production. It is different from the non-hydrolyzing enzymes for bacterial cell wall biosynthesis, and it is feedback inhibited by the downstream product CMP-Neu5Ac. Being a key enzyme that catalyzes the rate-limiting step of sialic acid biosynthesis, GNE plays an important role in regulation of cell-surface sialyation level by binding to the downstream product CMP-Neu5Ac. Regulation of cell-surface sialyation level by binding to the downstream product CMP-Neu5Ac. The feedback inhibition is highly positively cooperative and it does not affect the ManNAc kinase activity. By mediating cell-cell recognition, sialic acids are important in the development of nervous system. Structure and biosynthesis of sialic acid, overview | Homo sapiens |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
65.4 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant mutant S302A, pH 7.5, 37°C | Homo sapiens | |
152 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant mutant D112A, pH 7.5, 37°C | Homo sapiens | |
356.5 | - |
UDP-N-acetyl-alpha-D-glucosamine | recombinant wild-type enzyme, pH 7.5, 37°C | Homo sapiens |