N-glycosyltransferase

Last updated
Glycosyl transferase family 41
Identifiers
SymbolGT41
Pfam PF13844
CAZy GT41
Available protein structures:
Pfam   structures / ECOD  
PDB RCSB PDB; PDBe; PDBj
PDBsum structure summary

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.

Contents

Such enzymes have been found in the bacteria Actinobacillus pleuropneumoniae (whose N-glycosyltransferase is the best researched member of this enzyme family) and Haemophilus influenzae , and later in other bacterial species such as Escherichia coli . N-glycosyltransferases usually target adhesin proteins, which are involved in the attachment of bacterial cells to epithelia (in pathogenic bacteria); glycosylation is important for the stability and function of the adhesins.

History and definition

N-glycosyltransferase activity was first discovered in 2003 by St. Geme et al. in Haemophilus influenzae [1] and identified as a novel type of glycosyltransferase in 2010. [2] The Actinobacillus pleuropneumoniae N-glycosyltransferase is the best researched enzyme of this family. [3] [4] Initially, protein glycosylation was considered to be a purely eukaryotic process [5] before such processes were discovered in prokaryotes, including N-glycosyltransferases. [3]

Biochemistry

N-glycosyltransferases are an unusual [lower-alpha 1] type of glycosyltransferase which joins single hexoses to the target protein. [6] [7] [4] Attachment of sugars to the nitrogen atom in an amide group — such as the amide group of an asparagine — requires an enzyme, as the electrons of the nitrogen are delocalized in a pi-electron system with the carbon of the amide. Several mechanisms have been proposed for the activation. Among these are a deprotonation of the amide, an interaction between a hydroxyl group in the substrate sequon with the amide [9] [10] (a theory which is supported by the fact that the glycosylation rates appear to increase with the basicity of the second amino acid in the sequon [11] ) and two interactions involving acidic amino acids in the enzyme with each hydrogen atom of the amide group. This mechanism is supported by x-ray structures and biochemical information about glycosylation processes; the interaction breaks the delocalization and allows the electrons of the nitrogen to perform a nucleophilic attack on the sugar substrate. [8]

N-glycosyltransferases from Actinobacillus pleuropneumoniae [12] and Haemophilus influenzae use an asparagine-amino acid other than proline-serine or threonine sequences as target sequences, the same sequence used by oligosaccharyltransferases. [13] [14] The glutamine-469 residue in the Actinobacillus pleuropneumoniaeN-glycosyltransferase and its homologues in other N-glycosyltransferases is important for the selectivity of the enzyme. [15] The enzyme activity is further influenced by the amino acids around the sequon, with beta-loop structures especially important. [16] At least the Actinobacillus pleuropneumoniae N-glycosyltransferase can also hydrolyze sugar-nucleotides in the absence of a substrate, [17] a pattern frequently observed in glycosyltransferases, [18] and some N-glycosyltransferases can attach additional hexoses on oxygen atoms of the protein-linked hexose. [7] N-glycosylation by Actinobacillus pleuropneumoniae HMW1C does not require metals, [12] consistent with observations made on other GT41 family glycosyltransferases [19] and a distinction from oligosaccharyltransferases. [12]

Classification

Structurally N-glycosyltransferases belong to the GT41 family of glycosyltransferases and resemble protein O-GlcNAc transferase, an eukaryotic enzyme with various nuclear, mitochondrial and cytosolic targets. [8] Regular N-linked oligosaccharyltransferases belong to a different protein family, STT3. [20] The Haemophilus influenzaeN-glycosyltransferase has domains with homologies to glutathione S-transferase and glycogen synthase. [21]

The N-glycosyltransferases are subdivided into two functional classes, the first (e.g several Yersinia , Escherichia coli and Burkholderia sp.) is linked to trimeric autotransporter adhesins and the second has enzymes genomically linked to ribosome and carbohydrate metabolism associated proteins (e.g Actinobacillus pleuropneumoniae , Haemophilus ducreyi and Kingella kingae ). [22]

Functions

N-linked glycosylation is an important process, especially in eukaryotes where over half of all proteins have N-linked sugars attached [13] and where it is the most common form of glycosylation. [23] The processes are also important in prokaryotes [13] and archaeans. [24] In animals for example protein processing in the endoplasmic reticulum and several functions of the immune system are dependent on glycosylation. [9] [lower-alpha 2]

The principal substrates of N-glycosyltransferases are adhesins. [8] Adhesins are proteins that are used to colonize a surface, often a mucosal surface in the case of pathogenic bacteria. [27] N-glycosyltransferase homologues have been found in pathogenic gammaproteobacteria, [28] such as Yersinia and other pasteurellaceae. [8] These homologues are very similar to the Actinobacillus pleuropneumoniae enzyme and can glycosylate the Haemophilus influenzae HMW1A adhesin. [29]

N-glycosyltransferases may be a novel glycoengineering tool, [30] considering that they do not require a lipid carrier to perform their function. [31] Glycosylation is important for the function of many proteins and the production of glycosylated proteins can be a challenge. [26] Potential uses of glycoengineering tools include the creation of vaccines against protein-bound polysaccharides. [32]

Examples

Notes

  1. ↑ Regular N-glycosyltransferases are oligosaccharide-transferring enzymes. [6] [7] [4] Even though both enzyme families attach sugars to nitrogen, the Haemophilus influenzae N-glycosyltransferase bears no similarity to the oligosaccharyltransferases [8] and appears to have evolved independently. [1]
  2. ↑ N-glycosylation typically involves the attachment of oligosaccharides to asparagine amino groups in proteins; [13] the asparagine is usually followed two amino acids later by a serine or a threonine. [25] The oligosaccharide in most cases is assembled on an isoprenoid as carrier, [24] with a variety of oligosaccharides used. [26]

Related Research Articles

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

Asparagine is an α-amino acid that is used in the biosynthesis of proteins. It contains an α-amino group, an α-carboxylic acid group, and a side chain carboxamide, classifying it as a polar, aliphatic amino acid. It is non-essential in humans, meaning the body can synthesize it. It is encoded by the codons AAU and AAC.

<span class="mw-page-title-main">Glycoprotein</span> Protein with oligosaccharide modifications

Glycoproteins are proteins which contain oligosaccharide chains covalently attached to amino acid side-chains. The carbohydrate is attached to the protein in a cotranslational or posttranslational modification. This process is known as glycosylation. Secreted extracellular proteins are often glycosylated.

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.

<i>Haemophilus influenzae</i> Species of bacterium

Haemophilus influenzae is a Gram-negative, non-motile, coccobacillary, facultatively anaerobic, capnophilic pathogenic bacterium of the family Pasteurellaceae. The bacteria are mesophilic and grow best at temperatures between 35 and 37 °C.

<span class="mw-page-title-main">Pasteurellaceae</span> Family of bacteria

The Pasteurellaceae comprise a large family of Gram-negative bacteria. Most members live as commensals on mucosal surfaces of birds and mammals, especially in the upper respiratory tract. Pasteurellaceae are typically rod-shaped, and are a notable group of facultative anaerobes. Their biochemical characteristics can be distinguished from the related Enterobacteriaceae by the presence of oxidase, and from most other similar bacteria by the absence of flagella.

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

Oligosaccharyltransferase or OST (EC 2.4.1.119) is a membrane protein complex that transfers a 14-sugar oligosaccharide from dolichol to nascent protein. It is a type of glycosyltransferase. The sugar Glc3Man9GlcNAc2 (where Glc=Glucose, Man=Mannose, and GlcNAc=N-acetylglucosamine) is attached to an asparagine (Asn) residue in the sequence Asn-X-Ser or Asn-X-Thr where X is any amino acid except proline. This sequence is called a glycosylation sequon. The reaction catalyzed by OST is the central step in the N-linked glycosylation pathway.

<span class="mw-page-title-main">Glycosyltransferase</span> Class of enzymes that catalyze the transfer of glycosyl groups to an acceptor

Glycosyltransferases are enzymes that establish natural glycosidic linkages. They catalyze the transfer of saccharide moieties from an activated nucleotide sugar to a nucleophilic glycosyl acceptor molecule, the nucleophile of which can be oxygen- carbon-, nitrogen-, or sulfur-based.

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

Ribophorins are dome shaped transmembrane glycoproteins which are located in the membrane of the rough endoplasmic reticulum, but are absent in the membrane of the smooth endoplasmic reticulum. There are two types of ribophorines: ribophorin I and II. These act in the protein complex oligosaccharyltransferase (OST) as two different subunits of the named complex. Ribophorin I and II are only present in eukaryote cells.

In enzymology, a dolichyl-diphosphooligosaccharide–protein glycotransferase is an enzyme that catalyzes the chemical reaction

In enzymology, a protein N-acetylglucosaminyltransferase is an enzyme that catalyzes the chemical reaction

<span class="mw-page-title-main">ALG2</span> Protein-coding gene in the species Homo sapiens

Alpha-1,3/1,6-mannosyltransferase ALG2 is an enzyme that is encoded by the ALG2 gene. Mutations in the human gene are associated with congenital defects in glycosylation The protein encoded by the ALG2 gene belongs to two classes of enzymes: GDP-Man:Man1GlcNAc2-PP-dolichol alpha-1,3-mannosyltransferase and GDP-Man:Man2GlcNAc2-PP-dolichol alpha-1,6-mannosyltransferase.

<span class="mw-page-title-main">RPN2</span> Protein-coding gene in the species Homo sapiens

Dolichyl-diphosphooligosaccharide—protein glycosyltransferase subunit 2, also called ribophorin ǁ is an enzyme that in humans is encoded by the RPN2 gene.

<span class="mw-page-title-main">ALG8</span> Protein-coding gene in the species Homo sapiens

Probable dolichyl pyrophosphate Glc1Man9GlcNAc2 alpha-1,3-glucosyltransferase is an enzyme that in humans is encoded by the ALG8 gene.

<span class="mw-page-title-main">ALG12</span> Enzyme-coding gene in humans

Dolichyl-P-Man:Man(7)GlcNAc(2)-PP-dolichyl-alpha-1,6-mannosyltransferase is an enzyme that in humans is encoded by the ALG12 gene.

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

Dolichyl-diphosphooligosaccharide—protein glycosyltransferase 48 kDa subunit is an enzyme that in humans is encoded by the DDOST gene.

<span class="mw-page-title-main">ALG1</span> Protein-coding gene in the species Homo sapiens

Chitobiosyldiphosphodolichol beta-mannosyltransferase is an enzyme that is encoded by ALG1 whose structure and function has been conserved from lower to higher organisms.

<i>N</i>-linked glycosylation Attachment of an oligosaccharide to a nitrogen atom

N-linked glycosylation, is the attachment of an oligosaccharide, a carbohydrate consisting of several sugar molecules, sometimes also referred to as glycan, to a nitrogen atom, in a process called N-glycosylation, studied in biochemistry. The resulting protein is called an N-linked glycan, or simply an N-glycan.

<span class="mw-page-title-main">Trimeric autotransporter adhesin</span> Proteins found on the outer membrane of Gram-negative bacteria

In molecular biology, trimeric autotransporter adhesins (TAAs), are proteins found on the outer membrane of Gram-negative bacteria. Bacteria use TAAs in order to infect their host cells via a process called cell adhesion. TAAs also go by another name, oligomeric coiled-coil adhesins, which is shortened to OCAs. In essence, they are virulence factors, factors that make the bacteria harmful and infective to the host organism.

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.

The Nicotinamide Ribonucleoside (NR) Uptake Permease (PnuC) Family is a family of transmembrane transporters that is part of the TOG superfamily. Close PnuC homologues are found in a wide range of Gram-negative and Gram-positive bacteria, archaea and eukaryotes.

References

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  2. ↑ Choi et al. 2010, p. 2.
  3. 1 2 Song et al. 2017, p. 8856.
  4. 1 2 3 Naegeli & Aebi 2015, p. 11.
  5. ↑ Nothaft & Szymanski 2013, p. 6912.
  6. 1 2 3 4 Grass S, Lichti CF, Townsend RR, Gross J, St Geme JW (May 2010). "The Haemophilus influenzae HMW1C protein is a glycosyltransferase that transfers hexose residues to asparagine sites in the HMW1 adhesin". PLOS Pathogens. 6 (5): e1000919. doi: 10.1371/journal.ppat.1000919 . PMC   2877744 . PMID   20523900.
  7. 1 2 3 4 Gawthorne et al. 2014, p. 633.
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  9. 1 2 Naegeli et al. 2014, p. 24521.
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  11. ↑ Bause & Legler 1981, p. 644.
  12. 1 2 3 4 5 6 7 Schwarz et al. 2011, p. 35273.
  13. 1 2 3 4 5 6 7 Schwarz et al. 2011, p. 35267.
  14. 1 2 Piniello, Macías-León & Miyazaki 2023, p. 2.
  15. ↑ Song et al. 2017, p. 8861.
  16. ↑ Bause E (February 1983). "Structural requirements of N-glycosylation of proteins. Studies with proline peptides as conformational probes". The Biochemical Journal. 209 (2): 331–336. doi:10.1042/bj2090331. PMC   1154098 . PMID   6847620.
  17. 1 2 Naegeli et al. 2014, p. 24524.
  18. ↑ Naegeli et al. 2014, p. 24530.
  19. ↑ Choi et al. 2010, p. 7.
  20. 1 2 Naegeli et al. 2014, p. 2171.
  21. ↑ Kawai et al. 2011, p. 38553.
  22. 1 2 McCann & St Geme 2014, p. 2.
  23. 1 2 Choi et al. 2010, p. 1.
  24. 1 2 3 4 Naegeli et al. 2014, p. 2170.
  25. ↑ Bause & Legler 1981, p. 639.
  26. 1 2 Naegeli & Aebi 2015, p. 4.
  27. ↑ Grass et al. 2003, p. 737.
  28. ↑ Schwarz et al. 2011, p. 35269.
  29. ↑ Gawthorne et al. 2014, p. 636.
  30. 1 2 Song et al. 2017, p. 8857.
  31. 1 2 3 McCann & St Geme 2014, p. 3.
  32. ↑ Naegeli & Aebi 2015, p. 12.
  33. ↑ Naegeli et al. 2014, p. 2172.
  34. ↑ Keys TG, Wetter M, Hang I, Rutschmann C, Russo S, Mally M, et al. (November 2017). "A biosynthetic route for polysialylating proteins in Escherichia coli". Metabolic Engineering. 44: 293–301. doi: 10.1016/j.ymben.2017.10.012 . PMID   29101090.
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  36. 1 2 Naegeli et al. 2014, p. 2178.
  37. 1 2 Naegeli et al. 2014, p. 24531.
  38. ↑ Gawthorne et al. 2014, p. 634.
  39. ↑ Kawai et al. 2011, p. 38547.
  40. ↑ Kawai et al. 2011, p. 38549,38550.
  41. ↑ Cuccui et al. 2017, p. 2.
  42. ↑ Cuccui et al. 2017, p. 10.
  43. 1 2 3 Rempe et al. 2015, p. 5.
  44. 1 2 Rempe et al. 2015, p. 4.
  45. 1 2 3 McCann & St Geme 2014, p. 1.
  46. ↑ Rempe et al. 2015, p. 2.
  47. ↑ Gawthorne et al. 2014, p. 637,638.
  48. ↑ Grass et al. 2003, p. 742.
  49. ↑ Kawai et al. 2011, p. 38546.
  50. 1 2 Valguarnera E, Kinsella RL, Feldman MF (August 2016). "Sugar and Spice Make Bacteria Not Nice: Protein Glycosylation and Its Influence in Pathogenesis". Journal of Molecular Biology. 428 (16): 3206–3220. doi:10.1016/j.jmb.2016.04.013. PMID   27107636.
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  52. ↑ Rempe et al. 2015, p. 6.

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