Abstract
Asparagine-linked glycosylation, also known as N-linked glycosylation is an essential and highly conserved post-translational protein modification that occurs in all three domains of life. This modification is essential for specific molecular recognition, protein folding, sorting in the endoplasmic reticulum, cell–cell communication, and stability. Defects in N-linked glycosylation results in a class of inherited diseases known as congenital disorders of glycosylation (CDG). N-linked glycosylation occurs in the endoplasmic reticulum (ER) lumen by a membrane associated enzyme complex called the oligosaccharyltransferase (OST). In the central step of this reaction, an oligosaccharide group is transferred from a lipid-linked dolichol pyrophosphate donor to the acceptor substrate, the side chain of a specific asparagine residue of a newly synthesized protein. The prokaryotic OST enzyme consists of a single polypeptide chain, also known as single subunit OST or ssOST. In contrast, the eukaryotic OST is a complex of multiple non-identical subunits. In this review, we will discuss the biochemical and structural characterization of the prokaryotic, yeast, and mammalian OST enzymes. This review explains the most recent high-resolution structures of OST determined thus far and the mechanistic implication of N-linked glycosylation throughout all domains of life. It has been shown that the ssOST enzyme, AglB protein of the archaeon Archaeoglobus fulgidus, and the PglB protein of the bacterium Campylobactor lari are structurally and functionally similar to the catalytic Stt3 subunit of the eukaryotic OST enzyme complex. Yeast OST enzyme complex contains a single Stt3 subunit, whereas the human OST complex is formed with either STT3A or STT3B, two paralogues of Stt3. Both human OST complexes, OST-A (with STT3A) and OST-B (containing STT3B), are involved in the N-linked glycosylation of proteins in the ER. The cryo-EM structures of both human OST-A and OST-B complexes were reported recently. An acceptor peptide and a donor substrate (dolichylphosphate) were observed to be bound to the OST-B complex whereas only dolichylphosphate was bound to the OST-A complex suggesting disparate affinities of two OST complexes for the acceptor substrates. However, we still lack an understanding of the independent role of each eukaryotic OST subunit in N-linked glycosylation or in the stabilization of the enzyme complex. Discerning the role of each subunit through structure and function studies will potentially reveal the mechanistic details of N-linked glycosylation in higher organisms. Thus, getting an insight into the requirement of multiple non-identical subunits in the N-linked glycosylation process in eukaryotes poses an important future goal.
Highlights
Stt3 protein is the catalytic subunit in yeast OST, which is homologous to the single subunit OST enzyme, archaeal glycosylation B (AglB) in archaea, and protein glycosylation B (PglB) in bacteria [15]
The metazoan OST subunits have been identified and cloned. All of these protein subunits have homologs in the yeast OST complex [20] as shown in Table 1: ribophorin I is the homolog of yeast Ost1, DAD1 correspond to yeast Ost2, OST 4-kDa subunit (OST4) to Ost4, ribophorin II to yeast Swp1, tumor suppressor candidate 3 (TUSC3)/magnesium transporter protein 1 (MAGT1) to yeast Ost3/Ost6, transmembrane protein 258 (TMEM258) to yeast Ost5, OST 48-kDa subunit (OST48) to yeast Wbp1, and STT3A/STT3B to yeast Stt3 [21]
It is proposed that unlike the bacterial linked oligosaccharide (LLO), the yeast LLO is too large to dive under the disordered external loop5 (EL5); it enters the catalytic site via the gap between TMH8 and TMH9 of Stt3 [30]
Summary
N-linked glycosylation was originally believed to take place only in eukaryotic organisms until the discovery of alkali-sensitive glycoproteins extracted from the cell surface in archaea, Halobacterium [41,42,43,44,45]. The above oligosaccharides are shorter and not branched, the N-linked glycan of Pyrobaculum calidifontis has high mannose content with branching [48] while that of Archaeoglobus fulgidus has high hexose content with branching [49]. This evidence suggests a wide diversity in the N-glycan structures of the LLO donor substrates for the AglB enzyme dolichol is the common carrier [42,44,49]. The PglB enzyme in eubacteria transfers the donor substrate, a preassembled heptasaccharide attached to undecaprenyl pyrophosphate (UndPP-heptasaccharide) to a wide array of target proteins at selected asparagine residues in the consensus sequon
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