This difference makes for a protein with high glycan meta-heterogeneity, indicating selective regulation and highlighting potential differences in structurefunction relationships. == Examples of reported meta-heterogeneity in protein glycosylation WS3 == There is likely no better example to start with than immunoglobulin G (IgG), perhaps the most-studied glycoprotein in terms of glycosylation to date. higher level of glycan regulation: the variation in glycosylation across multiple sites of a given protein. Rabbit polyclonal to TIE1 We provide literature examples of extensive meta-heterogeneity on relevant proteins such as antibodies, erythropoietin, myeloperoxidase, and a number of serum and plasma proteins. Furthermore, we postulate around the possible biological reasons and causes behind the intriguing meta-heterogeneity observed in glycoproteins. Keywords:Meta-heterogeneity, Micro-heterogeneity, Macro-heterogeneity, Glycan, Glycosylation, Proteoforms, Glycoproteoforms, Plasma/serum glycoproteins, Immunoglobulins, Acute phase proteins Abbreviations:ApoB-100, apolipoprotein B-100; CID, collision-induced dissociation; ECD, electron-capture dissociation; EPO, erythropoietin; ETD, electron-transfer dissociation; Fab, fragment antigen-binding; HCD, higher-energy collision dissociation; MPO, myeloperoxidase;O-Man,O-mannosylation == Graphical Abstract == == Highlights == Micro- and macro-heterogeneity describe variation and occupancy of glycan sites. We propose meta-heterogeneity: glycan variation across multiple sites of a protein. Many glycoproteins exhibit meta-heterogeneity, including immunoglobulin G. Study of meta-heterogeneity will be critical to understand glycoprotein function. == In Brief == Diversity in protein glycosylation can be described in terms of micro-heterogeneity and macro-heterogeneity, respectively, referring to the variation and occupancy of glycans at a given glycosylation site. However, these terms are not sufficient to describe a higher level of regulation when proteins are multiply glycosylated. For this, we propose the term meta-heterogeneity: variation in glycosylation across multiple sites of a given glycoprotein. In this review, we describe several remarkable examples of glycoprotein meta-heterogeneity and underline the need for its investigation. Glycosylation is usually by far the most abundant posttranslational protein modification encountered in nature. Glycosylation plays critical roles in all facets of health and disease, and slight changes in the structure of the glycan attached to the polypeptide backbone can have a dramatic biological effect (1,2,3). Although often simply regarded as a uniform modification, it actually encompasses dozens of different types of glycosylation with new ones discovered regularly (Fig. 1) (4,5,6,7). == Fig. 1. == Overview of the glycosylation features discussed in this review that contribute to meta-heterogeneity. Glycan compositions were chosen to represent a small and a large variant within the given group (e.g., high-mannose). Tn, Core 1, and Core 2O-glycosylation represent only a selection of the large group of possibleO-glycan core structures, as the compositions themselves can be much larger.O-Acetylation has been positioned at a triantennary glycan but may in theory occur at any sialic acid (N-acetylneuraminic acid orN-glycolylneuraminic acid). Perhaps the most well-known and certainly the most studied type of glycosylation refers toN-linked protein glycosylation, orN-glycosylation. Here, a single uniform precursor consisting of twoN-acetylglucosamines WS3 (GlcNAc), nine mannoses (Man) and WS3 three glucoses (Glc), in short Glc3Man9GlcNAc2, is usually transferred cotranslationally to a side chain of asparagine in an Asn-Xxx-Ser/Thr sequon (where Xxx can be any amino acid besides proline). In addition,N-glycans may in some cases be transferred to an Asn-Xxx-Cys/Val sequon, but this appears to result in relatively low degrees of occupancy. A second widely encountered type of glycosylation is the so-called mucin-typeO-glycosylation, in which a singleO-GalNAc residue is usually transferred to a hydroxyl group of threonine, serine, or tyrosine (8,9,10,11). UnlikeN-glycosylation, in which a single precursor is usually transferred to the polypeptide backbone, initiation ofO-glycosylation can be catalyzed by at least 20 distinctO-GalNAc transferases, which have both overlapping and differential substrate preferences (12,13,14). InitiatingO-GalNAcs can also be further extended to create more elaboratedO-glycan structures, common examples including Core 1 (Gal1,3-GalNAc) and Core 2 (Gal1,3-[GlcNAc1,6-]GalNAc) typeO-glycans.O-Glycans mainly serve to stabilize the protein structure and protect against proteolytic cleavage (15), but they are also implicated in various pathophysiological states such as IgA nephropathy (16), Tn-syndrome (17), impaired leukocyte recruitment (18), and tumorigenesis (19). In addition to the mucin typeO-glycosylation, there are also numerous other classes ofO-glycosylation that have received so far less attention (9,20). Examples includeO-fucosylation (21,22,23) andO-glucosylation (24,25,26) found on epidermal growth factor-like and thrombospondin repeat domains,O-galactosylation (27,28) on collagen domain name hydroxylysines, andO-mannosylation (O-Man) mainly associated with the alpha-dystroglycan and cadherins (29). Although the aforementioned protein modifications are never found in the nucleocytoplasmic space,O-GlcNAcylation, catalyzed by theO-GlcNAc transferase, can be found in nuclear, cytoplasmic, and mitochondrial compartments (30). In contrast to the other types of modifications,O-GlcNAcylation is usually a dynamic modification that is thought to serve,e.g., as a nutrient sensor regulating transcription, signaling, mitochondrial activity, and cytoskeletal functions (31). Similar to the nucleocytoplasmicO-GlcNAcylation that is widespread in.