Glycosyltransferase

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Most glycosyltransferase enzymes form one of two folds: GT-A or GT-B Glycosyltransferase Folds.png
Most glycosyltransferase enzymes form one of two folds: GT-A or GT-B

Glycosyltransferases (GTFs, Gtfs) are enzymes (EC 2.4) that establish natural glycosidic linkages. They catalyze the transfer of saccharide moieties from an activated nucleotide sugar (also known as the "glycosyl donor") to a nucleophilic glycosyl acceptor molecule, the nucleophile of which can be oxygen- carbon-, nitrogen-, or sulfur-based. [1]

Contents

The result of glycosyl transfer can be a carbohydrate, glycoside, oligosaccharide, or a polysaccharide. Some glycosyltransferases catalyse transfer to inorganic phosphate or water. Glycosyl transfer can also occur to protein residues, usually to tyrosine, serine, or threonine to give O-linked glycoproteins, or to asparagine to give N-linked glycoproteins. Mannosyl groups may be transferred to tryptophan to generate C-mannosyl tryptophan, which is relatively abundant in eukaryotes. Transferases may also use lipids as an acceptor, forming glycolipids, and even use lipid-linked sugar phosphate donors, such as dolichol phosphates in eukaryotic organism, or undecaprenyl phosphate in bacteria.

Glycosyltransferases that use sugar nucleotide donors are Leloir enzymes, after Luis F. Leloir, the scientist who discovered the first sugar nucleotide and who received the 1970 Nobel Prize in Chemistry for his work on carbohydrate metabolism. Glycosyltransferases that use non-nucleotide donors such as dolichol or polyprenol pyrophosphate are non-Leloir glycosyltransferases.

Mammals use only 9 sugar nucleotide donors for glycosyltransferases: [2] UDP-glucose, UDP-galactose, UDP-GlcNAc, UDP-GalNAc, UDP-xylose, UDP-glucuronic acid, GDP-mannose, GDP-fucose, and CMP-sialic acid. The phosphate(s) of these donor molecules are usually coordinated by divalent cations such as manganese, however metal independent enzymes exist.

Many glycosyltransferases are single-pass transmembrane proteins, and they are usually anchored to membranes of Golgi apparatus [3]

Mechanism

Glycosyltransferase mechanisms.png

Glycosyltransferases can be segregated into "retaining" or "inverting" enzymes according to whether the stereochemistry of the donor's anomeric bond is retained (α→α) or inverted (α→β) during the transfer. The inverting mechanism is straightforward, requiring a single nucleophilic attack from the accepting atom to invert stereochemistry.

The retaining mechanism has been a matter of debate, but there exists strong evidence against a double displacement mechanism (which would cause two inversions about the anomeric carbon for a net retention of stereochemistry) or a dissociative mechanism (a prevalent variant of which was known as SNi). An "orthogonal associative" mechanism has been proposed which, akin to the inverting enzymes, requires only a single nucleophilic attack from an acceptor from a non-linear angle (as observed in many crystal structures) to achieve anomer retention. [4]

Reaction reversibility

The recent discovery of the reversibility of many reactions catalyzed by inverting glycosyltransferases served as a paradigm shift in the field and raises questions regarding the designation of sugar nucleotides as 'activated' donors. [5] [6] [7] [8] [9]

Classification by sequence

Sequence-based classification methods have proven to be a powerful way of generating hypotheses for protein function based on sequence alignment to related proteins. The carbohydrate-active enzyme database presents a sequence-based classification of glycosyltransferases into over 90 families. [10] The same three-dimensional fold is expected to occur within each of the families. [11]

Structure

In contrast to the diversity of 3D structures observed for glycoside hydrolases, glycosyltransferase have a much smaller range of structures. [12] [13] In fact, according to the Structural Classification of Proteins database, only three different folds have been observed for glycosyltransferases [14] Very recently, a new glycosyltransferase fold was identified for the glycosyltransferases involved in the biosynthesis of the NAG-NAM polymer backbone of peptidoglycan. [15]

Inhibitors

Many inhibitors of glycosyltransferases are known. Some of these are natural products, such as moenomycin, an inhibitor of peptidoglycan glycosyltransferases, the nikkomycins, inhibitors of chitin synthase, and the echinocandins, inhibitors of fungal β-1,3-glucan synthases. Some glycosyltransferase inhibitors are of use as drugs or antibiotics. Moenomycin is used in animal feed as a growth promoter. Caspofungin has been developed from the echinocandins and is in use as an antifungal agent. Ethambutol is an inhibitor of mycobacterial arabinotransferases and is used for the treatment of tuberculosis. Lufenuron is an inhibitor of insect chitin syntheses and is used to control fleas in animals. Imidazolium-based synthetic inhibitors of glycosyltransferases have been designed for use as antimicrobial and antiseptic agents. [16]

Determinant of blood type

Glycosyltransferase family 6
Identifiers
SymbolGT6
Pfam PF03414
InterPro IPR005076
OPM superfamily 199
OPM protein 2rj6
Membranome 468
Available protein structures:
Pfam   structures / ECOD  
PDB RCSB PDB; PDBe; PDBj
PDBsum structure summary

The ABO blood group system is determined by what type of glycosyltransferases are expressed in the body.

The ABO gene locus expressing the glycosyltransferases has three main allelic forms: A, B, and O. The A allele encodes 1-3-N-acetylgalactosaminyltransferase that bonds α-N-acetylgalactosamine to D-galactose end of H antigen, producing the A antigen. The B allele encodes 1-3-galactosyltransferase that joins α-D-galactose bonded to D-galactose end of H antigen, creating the B antigen. In case of O allele the exon 6 contains a deletion that results in a loss of enzymatic activity. The O allele differs slightly from the A allele by deletion of a single nucleotide - Guanine at position 261. The deletion causes a frameshift and results in translation of an almost entirely different protein that lacks enzymatic activity. This results in H antigen remaining unchanged in case of O groups.

The combination of glycosyltransferases by both alleles present in each person determines whether there is an AB, A, B or O blood type.

Uses

Glycosyltransferases have been widely used in both the targeted synthesis of specific glycoconjugates as well as the synthesis of differentially glycosylated libraries of drugs, biological probes or natural products in the context of drug discovery and drug development (a process known as glycorandomization). [17] Suitable enzymes can be isolated from natural sources or produced recombinantly. As an alternative, whole cell-based systems using either endogenous glycosyl donors or cell-based systems containing cloned and expressed systems for synthesis of glycosyl donors have been developed. In cell-free approaches, the large-scale application of glycosyltransferases for glycoconjugate synthesis has required access to large quantities of the glycosyl donors. On the flip-side, nucleotide recycling systems that allow the resynthesis of glycosyl donors from the released nucleotide have been developed. The nucleotide recycling approach has a further benefit of reducing the amount of nucleotide formed as a by-product, thereby reducing the amount of inhibition caused to the glycosyltransferase of interest – a commonly observed feature of the nucleotide byproduct.

See also

Related Research Articles

<span class="mw-page-title-main">Galactose</span> Monosaccharide sugar

Galactose, sometimes abbreviated Gal, is a monosaccharide sugar that is about as sweet as glucose, and about 65% as sweet as sucrose. It is an aldohexose and a C-4 epimer of glucose. A galactose molecule linked with a glucose molecule forms a lactose molecule.

A glycosidic bond or glycosidic linkage is a type of ether bond that joins a carbohydrate (sugar) molecule to another group, which may or may not be another carbohydrate.

Glycosylation is the reaction in which a carbohydrate, i.e. a glycosyl donor, is attached to a hydroxyl or other functional group of another molecule in order to form a glycoconjugate. In biology, glycosylation usually refers to an enzyme-catalysed reaction, whereas glycation may refer to a non-enzymatic reaction.

<span class="mw-page-title-main">Galactokinase</span> Enzyme

Galactokinase is an enzyme (phosphotransferase) that facilitates the phosphorylation of α-D-galactose to galactose 1-phosphate at the expense of one molecule of ATP. Galactokinase catalyzes the second step of the Leloir pathway, a metabolic pathway found in most organisms for the catabolism of α-D-galactose to glucose 1-phosphate. First isolated from mammalian liver, galactokinase has been studied extensively in yeast, archaea, plants, and humans.

<span class="mw-page-title-main">Glycolipid</span> Class of chemical compounds

Glycolipids are lipids with a carbohydrate attached by a glycosidic (covalent) bond. Their role is to maintain the stability of the cell membrane and to facilitate cellular recognition, which is crucial to the immune response and in the connections that allow cells to connect to one another to form tissues. Glycolipids are found on the surface of all eukaryotic cell membranes, where they extend from the phospholipid bilayer into the extracellular environment.

An Endoglycosidase is an enzyme that releases oligosaccharides from glycoproteins or glycolipids. It may also cleave polysaccharide chains between residues that are not the terminal residue, although releasing oligosaccharides from conjugated protein and lipid molecules is more common.

<span class="mw-page-title-main">Sialyltransferase</span> Class of enzymes

Sialyltransferases are enzymes that transfer sialic acid to nascent oligosaccharide. Each sialyltransferase is specific for a particular sugar substrate. Sialyltransferases add sialic acid to the terminal portions of the sialylated glycolipids (gangliosides) or to the N- or O-linked sugar chains of glycoproteins.

<span class="mw-page-title-main">Calicheamicin</span> Chemical compound

The calicheamicins are a class of enediyne antitumor antibiotics derived from the bacterium Micromonospora echinospora, with calicheamicin γ1 being the most notable. It was isolated originally in the mid-1980s from the chalky soil, or "caliche pits", located in Kerrville, Texas. The sample was collected by a scientist working for Lederle Labs. It is extremely toxic to all cells and, in 2000, a CD33 antigen-targeted immunoconjugate N-acetyl dimethyl hydrazide calicheamicin was developed and marketed as targeted therapy against the non-solid tumor cancer acute myeloid leukemia (AML). A second calicheamicin-linked monoclonal antibody, inotuzumab ozogamicin, an anti-CD22-directed antibody-drug conjugate, was approved by the U.S. Food and Drug Administration on August 17, 2017, for use in the treatment of adults with relapsed or refractory B-cell precursor acute lymphoblastic leukemia. Calicheamicin γ1 and the related enediyne esperamicin are the two of the most potent antitumor agents known.

<span class="mw-page-title-main">UTP—glucose-1-phosphate uridylyltransferase</span> Class of enzymes

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<span class="mw-page-title-main">Galactosyltransferase</span> Class of enzymes

Galactosyltransferase is a type of glycosyltransferase which catalyzes the transfer of galactose. An example is B-N-acetylglucosaminyl-glycopeptide b-1,4-galactosyltransferase.

<span class="mw-page-title-main">UDP-glucose 4-epimerase</span> Class of enzymes

The enzyme UDP-glucose 4-epimerase, 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. GALE tightly binds nicotinamide adenine dinucleotide (NAD+), a co-factor required for catalytic activity.

Nucleotide sugars are the activated forms of monosaccharides. Nucleotide sugars act as glycosyl donors in glycosylation reactions. Those reactions are catalyzed by a group of enzymes called glycosyltransferases.

<span class="mw-page-title-main">UDP-glucose—hexose-1-phosphate uridylyltransferase</span> Class of enzymes

In enzymology, an UDP-glucose—hexose-1-phosphate uridylyltransferase is an enzyme that catalyzes the chemical reaction

The gal operon is a prokaryotic operon, which encodes enzymes necessary for galactose metabolism. Repression of gene expression for this operon works via binding of repressor molecules to two operators. These repressors dimerize, creating a loop in the DNA. The loop as well as hindrance from the external operator prevent RNA polymerase from binding to the promoter, and thus prevent transcription. Additionally, since the metabolism of galactose in the cell is involved in both anabolic and catabolic pathways, a novel regulatory system using two promoters for differential repression has been identified and characterized within the context of the gal operon.

<span class="mw-page-title-main">ABO (gene)</span> Protein-coding gene in the species Homo sapiens

Histo-blood group ABO system transferase is an enzyme with glycosyltransferase activity, which is encoded by the ABO gene in humans. It is ubiquitously expressed in many tissues and cell types. ABO determines the ABO blood group of an individual by modifying the oligosaccharides on cell surface glycoproteins. Variations in the sequence of the protein between individuals determine the type of modification and the blood group. The ABO gene also contains one of 27 SNPs associated with increased risk of coronary artery disease.

Glycorandomization, is a drug discovery and drug development technology platform to enable the rapid diversification of bioactive small molecules, drug leads and/or approved drugs through the attachment of sugars. Initially developed as a facile method to manipulate carbohydrate substitutions of naturally occurring glycosides to afford the corresponding differentially glycosylated natural product libraries, glycorandomization applications have expanded to include both small molecules and even macromolecules (proteins). Also referred to as 'glycodiversification', glycorandomization has led to the discovery of new glycoside analogs which display improvements in potency, selectivity and/or ADMET as compared to the parent molecule.

O-linked glycosylation is the attachment of a sugar molecule to the oxygen atom of serine (Ser) or threonine (Thr) residues in a protein. O-glycosylation is a post-translational modification that occurs after the protein has been synthesised. In eukaryotes, it occurs in the endoplasmic reticulum, Golgi apparatus and occasionally in the cytoplasm; in prokaryotes, it occurs in the cytoplasm. Several different sugars can be added to the serine or threonine, and they affect the protein in different ways by changing protein stability and regulating protein activity. O-glycans, which are the sugars added to the serine or threonine, have numerous functions throughout the body, including trafficking of cells in the immune system, allowing recognition of foreign material, controlling cell metabolism and providing cartilage and tendon flexibility. Because of the many functions they have, changes in O-glycosylation are important in many diseases including cancer, diabetes and Alzheimer's. O-glycosylation occurs in all domains of life, including eukaryotes, archaea and a number of pathogenic bacteria including Burkholderia cenocepacia, Neisseria gonorrhoeae and Acinetobacter baumannii.

<span class="mw-page-title-main">Leloir pathway</span>

The Leloir pathway is a metabolic pathway for the catabolism of D-galactose. It is named after Luis Federico Leloir, who first described it.

Protein <i>O</i>-GlcNAc transferase Protein-coding gene in the species Homo sapiens

Protein O-GlcNAc transferase also known as OGT or O-linked N-acetylglucosaminyltransferase is an enzyme that in humans is encoded by the OGT gene. OGT catalyzes the addition of the O-GlcNAc post-translational modification to proteins.

N-glycosyltransferase is an enzyme in prokaryotes which transfers individual hexoses onto asparagine sidechains in substrate proteins, using a nucleotide-bound intermediary, within the cytoplasm. They are distinct from regular N-glycosylating enzymes, which are oligosaccharyltransferases that transfer pre-assembled oligosaccharides. Both enzyme families however target a shared amino acid sequence asparagine—-any amino acid except proline—serine or threonine (N–x–S/T), with some variations.

References

  1. ↑ Williams, GJ; Thorson, JS (2009). Natural product glycosyltransferases: properties and applications. Advances in Enzymology - and Related Areas of Molecular Biology. Vol. 76. pp. 55–119. doi:10.1002/9780470392881.ch2. ISBN   9780470392881. PMID   18990828.
  2. ↑ Etzler ME, Varki A, Cummings RL, Esko JD, Freeze HH, Hart GW, eds. (2008). Essentials of Glycobiology (2nd ed.). Plainview, N.Y: Cold Spring Harbor Laboratory Press. ISBN   978-0-87969-770-9.
  3. ↑ Transferases in Membranome database.
  4. ↑ Schuman B, Evans SV, Fyles TM (August 2013). "Geometric Attributes of Retaining Glycosyltransferase Enzymes Favor an Orthogonal Mechanism". PLOS ONE. 8 (8): e71077. Bibcode:2013PLoSO...871077S. doi: 10.1371/journal.pone.0071077 . PMC   3731257 . PMID   23936487.
  5. ↑ Zhang, C; Griffith, BR; Fu, Q; Albermann, C; Fu, X; Lee, IK; Li, L; Thorson, JS (1 September 2006). "Exploiting the reversibility of natural product glycosyltransferase-catalyzed reactions". Science. 313 (5791): 1291–4. Bibcode:2006Sci...313.1291Z. doi:10.1126/science.1130028. PMID   16946071. S2CID   38072017.
  6. ↑ Zhang, C; Albermann, C; Fu, X; Thorson, JS (27 December 2006). "The in vitro characterization of the iterative avermectin glycosyltransferase AveBI reveals reaction reversibility and sugar nucleotide flexibility". Journal of the American Chemical Society. 128 (51): 16420–1. doi:10.1021/ja065950k. PMID   17177349.
  7. ↑ Zhang, C; Fu, Q; Albermann, C; Li, L; Thorson, JS (5 March 2007). "The in vitro characterization of the erythronolide mycarosyltransferase EryBV and its utility in macrolide diversification". ChemBioChem. 8 (4): 385–90. doi:10.1002/cbic.200600509. PMID   17262863. S2CID   45058028.
  8. ↑ Zhang, C; Moretti, R; Jiang, J; Thorson, JS (13 October 2008). "The in vitro characterization of polyene glycosyltransferases AmphDI and NysDI". ChemBioChem. 9 (15): 2506–14. doi:10.1002/cbic.200800349. PMC   2947747 . PMID   18798210.
  9. ↑ Gantt, RW; Peltier-Pain, P; Cournoyer, WJ; Thorson, JS (21 August 2011). "Using simple donors to drive the equilibria of glycosyltransferase-catalyzed reactions". Nature Chemical Biology. 7 (10): 685–91. doi:10.1038/nchembio.638. PMC   3177962 . PMID   21857660.
  10. ↑ CAZypedia Glycosyltransferases
  11. ↑ "CAZy Glycosyl Transferase". Archived from the original on 2009-03-23. Retrieved 2009-08-25.
  12. ↑ Singh, S; Phillips GN, Jr; Thorson, JS (October 2012). "The structural biology of enzymes involved in natural product glycosylation". Natural Product Reports. 29 (10): 1201–37. doi:10.1039/c2np20039b. PMC   3627186 . PMID   22688446.
  13. ↑ Chang, A; Singh, S; Phillips GN, Jr; Thorson, JS (December 2011). "Glycosyltransferase structural biology and its role in the design of catalysts for glycosylation". Current Opinion in Biotechnology. 22 (6): 800–8. doi:10.1016/j.copbio.2011.04.013. PMC   3163058 . PMID   21592771.
  14. ↑ SCOP: Structural Classification of Proteins
  15. ↑ Lovering AL, de Castro LH, Lim D, Strynadka NC (March 2007). "Structural insight into the transglycosylation step of bacterial cell-wall biosynthesis". Science. 315 (5817): 1402–5. Bibcode:2007Sci...315.1402L. doi:10.1126/science.1136611. PMID   17347437. S2CID   41658295.
  16. ↑ Kocev, A; Melamed, J; Wang, S; Kong, X; Vlahakis, JZ; Xu, Y; Szarek, WA; Brockhausen, I (June 2020). "Inhibition of bacterial growth and galactosyltransferase activity of WbwC by α, ω-bis(3-alkyl-1H-imidazolium)alkane salts: Effect of varying carbon content". Bioorganic and Medicinal Chemistry. 28 (11): 115494. doi:10.1016/j.bmc.2020.115494. PMID 32312486.
  17. ↑ Gantt, RW; Peltier-Pain, P; Thorson, JS (October 2011). "Enzymatic methods for glyco(diversification/randomization) of drugs and small molecules". Natural Product Reports. 28 (11): 1811–53. doi:10.1039/c1np00045d. PMID   21901218.
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