UDP-glucose 4-epimerase
Class of enzymes From Wikipedia, the free encyclopedia
The enzyme UDP-glucose 4-epimerase (EC 5.1.3.2), also known as UDP-galactose 4-epimerase or GALE, is a homodimeric epimerase found in bacterial, fungal, plant, and mammalian cells. This enzyme performs the final step in the Leloir pathway of galactose metabolism, catalyzing the reversible conversion of UDP-galactose to UDP-glucose.[1] GALE tightly binds nicotinamide adenine dinucleotide (NAD+), a co-factor required for catalytic activity.[2]
| UDP-glucose 4-epimerase | |||||||||||||||||||||||||||||||||||||||||||||||||||
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| Aliases | UDPgalactose 4-epimerase4-epimeraseuridine diphosphate glucose 4-epimeraseUDPG-4-epimeraseUDP-galactose 4-epimeraseuridine diphosphoglucose epimeraseuridine diphospho-galactose-4-epimeraseUDP-D-galactose 4-epimeraseUDP-glucose epimeraseuridine diphosphoglucose 4-epimeraseuridine diphosphate galactose 4-epimerase | ||||||||||||||||||||||||||||||||||||||||||||||||||
| External IDs | GeneCards: ; OMA:- orthologs | ||||||||||||||||||||||||||||||||||||||||||||||||||
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| UDP-glucose 4-epimerase | |||||||||
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H. sapiens UDP-glucose 4-epimerase homodimer bound to NADH and UDP-glucose. Domains: N-terminal and C-terminal. | |||||||||
| Identifiers | |||||||||
| EC no. | 5.1.3.2 | ||||||||
| CAS no. | 9032-89-7 | ||||||||
| Databases | |||||||||
| IntEnz | IntEnz view | ||||||||
| BRENDA | BRENDA entry | ||||||||
| ExPASy | NiceZyme view | ||||||||
| KEGG | KEGG entry | ||||||||
| MetaCyc | metabolic pathway | ||||||||
| PRIAM | profile | ||||||||
| PDB structures | RCSB PDB PDBe PDBsum | ||||||||
| Gene Ontology | AmiGO / QuickGO | ||||||||
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| UDP-galactose-4-epimerase | |||||||
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 Human GALE bound to NAD+ and UDP-GlcNAc, with N- and C-terminal domains highlighted. Asn 207 contorts to accommodate UDP-GlcNAc within the active site. | |||||||
| Identifiers | |||||||
| Symbol | GALE | ||||||
| NCBI gene | 2582 | ||||||
| HGNC | 4116 | ||||||
| OMIM | 606953 | ||||||
| RefSeq | NM_000403 | ||||||
| UniProt | Q14376 | ||||||
| Other data | |||||||
| EC number | 5.1.3.2 | ||||||
| Locus | Chr. 1 p36-p35 | ||||||
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| NAD-dependent epimerase/dehydratase | |||||||||
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| Identifiers | |||||||||
| Symbol | ? | ||||||||
| Pfam | PF01370 | ||||||||
| InterPro | IPR001509 | ||||||||
| Membranome | 330 | ||||||||
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Additionally, human and some bacterial GALE isoforms reversibly catalyze the formation of UDP-N-acetylgalactosamine (UDP-GalNAc) from UDP-N-acetylglucosamine (UDP-GlcNAc) in the presence of NAD+, an initial step in glycoprotein or glycolipid synthesis.[3]
Historical significance
Dr. Luis Leloir deduced the role of GALE in galactose metabolism during his tenure at the Instituto de Investigaciones Bioquímicas del Fundación Campomar, initially terming the enzyme waldenase.[4] Dr. Leloir was awarded the 1970 Nobel Prize in Chemistry for his discovery of sugar nucleotides and their role in the biosynthesis of carbohydrates.[5]
Structure
Summarize
Perspective
GALE belongs to the short-chain dehydrogenase/reductase (SDR) superfamily of proteins.[6] This family is characterized by a conserved Tyr-X-X-X-Lys motif necessary for enzymatic activity; one or more Rossmann fold scaffolds; and the ability to bind NAD+.[6]
Tertiary structure
GALE structure has been resolved for a number of species, including E. coli[7] and humans.[8] GALE exists as a homodimer in various species.[8]
While subunit size varies from 68 amino acids (Enterococcus faecalis) to 564 amino acids (Rhodococcus jostii), a majority of GALE subunits cluster near 330 amino acids in length.[6] Each subunit contains two distinct domains. An N-terminal domain contains a 7-stranded parallel β-pleated sheet flanked by α-helices.[1] Paired Rossmann folds within this domain allow GALE to tightly bind one NAD+ cofactor per subunit.[2] A 6-stranded β-sheet and 5 α-helices comprise GALE's C-terminal domain.[1] C-terminal residues bind UDP, such that the subunit is responsible for correctly positioning UDP-glucose or UDP-galactose for catalysis.[1]
Active site
The cleft between GALE's N- and C-terminal domains constitutes the enzyme's active site. A conserved Tyr-X-X-X Lys motif is necessary for GALE catalytic activity; in humans, this motif is represented by Tyr 157-Gly-Lys-Ser-Lys 161,[6] while E. coli GALE contains Tyr 149-Gly-Lys-Ser-Lys 153.[8] The size and shape of GALE's active site varies across species, allowing for variable GALE substrate specificity.[3] Additionally, the conformation of the active site within a species-specific GALE is malleable; for instance, a bulky UDP-GlcNAc 2' N-acetyl group is accommodated within the human GALE active site by the rotation of the Asn 207 carboxamide side chain.[3]
| Residue | Function |
|---|---|
| Ala 216, Phe 218 | Anchor uracil ring to enzyme. |
| Asp 295 | Interacts with ribose 2' hydroxyl group. |
| Asn 179, Arg 231, Arg 292 | Interact with UDP phosphate groups. |
| Tyr 299, Asn 179 | Interact with galactose 2' hydroxyl or glucose 6' hydroxyl group; properly position sugar within active site. |
| Tyr 177, Phe 178 | Interact with galactose 3' hydroxyl or glucose 6' hydroxyl group; properly position sugar within active site. |
| Lys 153 | Lowers pKa of Tyr 149, allows for abstraction or donation of a hydrogen atom to or from the sugar 4' hydroxyl group. |
| Tyr 149 | Abstracts or donates a hydrogen atom to or from the sugar 4' hydroxyl group, catalyzing formation of 4-ketopyranose intermediate. |
Mechanism
Conversion of UDP-galactose to UDP-glucose
GALE inverts the configuration of the 4' hydroxyl group of UDP-galactose through a series of 4 steps. Upon binding UDP-galactose, a conserved tyrosine residue in the active site abstracts a proton from the 4' hydroxyl group.[7][10]
Concomitantly, the 4' hydride is added to the si-face of NAD+, generating NADH and a 4-ketopyranose intermediate.[1] The 4-ketopyranose intermediate rotates 180° about the pyrophosphoryl linkage between the glycosyl oxygen and β-phosphorus atom, presenting the opposite face of the ketopyranose intermediate to NADH.[10] Hydride transfer from NADH to this opposite face inverts the stereochemistry of the 4' center. The conserved tyrosine residue then donates its proton, regenerating the 4' hydroxyl group.[1]
Conversion of UDP-GlcNAc to UDP-GalNAc
Human and some bacterial GALE isoforms reversibly catalyze the conversion of UDP-GlcNAc to UDP-GalNAc through an identical mechanism, inverting the stereochemical configuration at the sugar's 4' hydroxyl group.[3][11]
Biological function
Summarize
Perspective
Galactose metabolism
No direct catabolic pathways exist for galactose metabolism. Galactose is therefore preferentially converted into glucose-1-phosphate, which may be shunted into glycolysis or the inositol synthesis pathway.[12]
GALE functions as one of four enzymes in the Leloir pathway of galactose conversion of glucose-1-phosphate. First, galactose mutarotase converts β-D-galactose to α-D-galactose.[1] Galactokinase then phosphorylates α-D-galactose at the 1' hydroxyl group, yielding galactose-1-phosphate.[1] In the third step, galactose-1-phosphate uridyltransferase catalyzes the reversible transfer of a UMP moiety from UDP-glucose to galactose-1-phosphate, generating UDP-galactose and glucose-1-phosphate.[1] In the final Leloir step, UDP-glucose is regenerated from UDP-galactose by GALE; UDP-glucose cycles back to the third step of the pathway.[1] As such, GALE regenerates a substrate necessary for continued Leloir pathway cycling.
The glucose-1-phosphate generated in step 3 of the Leloir pathway may be isomerized to glucose-6-phosphate by phosphoglucomutase. Glucose-6-phosphate readily enters glycolysis, leading to the production of ATP and pyruvate.[13] Furthermore, glucose-6-phosphate may be converted to inositol-1-phosphate by inositol-3-phosphate synthase, generating a precursor needed for inositol biosynthesis.[14]
UDP-GalNAc synthesis
Human and selected bacterial GALE isoforms bind UDP-GlcNAc, reversibly catalyzing its conversion to UDP-GalNAc. A family of glycosyltransferases known as UDP-N-acetylgalactosamine:polypeptide N-acetylgalactosamine transferases (ppGaNTases) transfers GalNAc from UDP-GalNAc to glycoprotein serine and threonine residues.[15] ppGaNTase-mediated glycosylation regulates protein sorting,[16][17][18][19][20] ligand signaling,[21][22][23] resistance to proteolytic attack,[24][25] and represents the first committed step in mucin biosynthesis.[15]
Role in disease
Human GALE deficiency or dysfunction results in Type III galactosemia, which may exist in a mild (peripheral) or more severe (generalized) form.[12]
References
Further reading
External links
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