(Glyco)Protein Folding Disorders
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The term “protein folding disorders” (PFD) was introduced for a small group of proteins that misfold after biosynthesis producing pathological symptoms, for example, by depositing in inappropriate places. Of these proteins, three can be normally or sometimes N-glycosylated hence the term “(glyco)protein folding disorders” (GFD). However one of these, the Aβ precursor glycoprotein providing the 40, 41, or 42 amino acid residue fragments called amyloid-β-protein (Aβ) that are found in the plaques of patients with Alzheimer’s disease (AD), is also O-GlcNAcylated and many other proteins can also be, which results in misfolding and tissue deposition.
The term “PFD” can also now be applied to many more proteins, as we find out more about their complicated life stories. For example, in Huntington’s Disease (HD), the addition at gene level of repeat CAG means that the protein can be biosynthesized with a long Gln tail. This is somewhat reminiscent of hemoglobin in sickle cell disease where a Glu to Val mutation results in a sticky end resulting in aggregation of hemoglobin and subsequent changing shape of the red blood cells. However, generally PFD relates to the amyloidoses (Omtvedt et al. 2000), a heterogeneous group of disorders in which normally soluble proteins aggregate to form insoluble amyloid deposits resistant to proteolysis.
The prion, the PRoteinacious INfectious particle that is the pathological reagent in the encephalopathies affecting many species, but including bovine spongiform encephalopathy (BSE) and, in humans, Creutzfeldt Jakob Disease (CJD);
Immunoglobulin light chains involved in systemic amyloidosis;
Alzheimer’s glycoprotein which is a transmembrane Type I glycoprotein that has N-glycans on the extracellular surface and O-GlcNAc (O-GlcNAcylation) on the cytoplasmically oriented side.
The nonspecific cases include many cytoplasmic proteins that have O-GlcNAcylation and the secreted (glyco)proteins that are modified by glycation. It could also be said that the many proteins that have O-linked glycosylation that is intimately involved in glycopeptide conformation and antigenicity could also rate as GFDs, but here we will stick to the three specific cases outlined above.
The GFD field really came alive (and defined) when the structure of the prion was elucidated and also it was shown that blockade of glycosylation promotes acquisition of scrapie-like properties by the prion protein in cultured cells (Lehmann and Harris 1997). The natural prion protein always has two sequons for N-glycosylation at or near (depending on the species) Asn 181 and 197. These can be and are glycosylated with hundreds of different glycoforms, and it is therefore difficult to see the importance of any one, but more interestingly there is preponderance for underglycosylation so that one can define three states having either one, both, or neither sites glycosylated. These can be seen, for example, by polyacrylamide gel electrophoresis as glycosylation gives a large additional molecular weight and the pattern of glycosylation occupancy defines different prion types (Wong et al. 2000). Another defining feature of the prion is that it is anchored in the cell membrane by a Glycosylphosphatidyl (GPI) moiety which can theoretically account for its infectivity as such a molecule may be able to move from cell membrane to cell membrane which is not allowed by integral transmembrane glycoproteins.
Alzheimer’s Disease (AD)
There are some similarities of the AD glycoprotein and the prion. The former is a transmembrane N-glycosylated glycoprotein where the transmembrane region has some sequence similarity with a section of (non-transmembrane) prion sequences (the prion is anchored not by this part, but by its GPI anchor) that is specifically cleaved by protease to give pathological peptides. Unlike the prion, N-glycosylation has not been implicated in the formation and deposition of amyloid-β-protein (Aβ) that are found in the plaques of patients with Alzheimer’s disease (AD); however the O-GlcNAcylation seems to be involved in pathogenesis. This is a common feature of many neuron-associated proteins, for example, neurofilaments, microtubule-associated proteins such as Tau, and clathrin assembly proteins. In AD, Tau is hyperphosphorylated causing it to dissociate and self-assemble into paired helical filaments which are the major components of neurofibrillary tangles in the brain (Wang et al. 1996). Phosphorylation is reciprocal with O-GlcNAcylation. O-GlcNAc modification of the clathrin assembly protein AP-3 is also reduced in AD, and this is associated with an increase in density of neurofibrillary tangle. A very large amount of glucose is converted to GlcNAc in neurons. Also cellular conversion is an essential prerequisite for the development of insulin-resistant or type 2 diabetes (Brooks et al. 2002). Advanced glycation end products (AGEs) contribute to amyloidosis in Alzheimer’s disease (Vitek et al. 1994) as has been proposed in several other pathologies such as involvement of β-2-microglobulin modified with AGEs in the pathogenesis of hemodialysis-associated amyloidosis (Miyata et al. 1994).
AL-amyloidosis is a fatal disease caused by deposition of Ig light chains in a fibrillar form (AL) in various organs. Hence this is also known as systemic amyloidosis by comparison to those primarily associated with the brain. AL-amyloidosis presents in patients with multiple myeloma or other plasma cell dyscrasias where there is an increase in production and secretion of oligoclonal or monoclonal immunoglobulin (Ig) components. However, only 10–15% of patients with multiple myeloma develop AL and the reason for this selectivity is unknown (Omtvedt et al. 2000). Approximately, 15% of circulating light chains from homogeneous Ig in patients with multiple myeloma have covalently attached oligosaccharides, and AL-associated proteins are four times more frequent than that reported for light chains from patients with myelomatosis without amyloidosis. Further, there is an unexpected preponderance of the glycosylation sequon in the framework regions of amyloid light chains versus the hypervariable regions (Omtvedt et al. 2000). A particular glycosylation pattern has been found with a bisecting GlcNAc (Omtvedt et al. 2000; Routier et al. 1998) which may be involved in tissue deposition. In addition, glycosylation may be important in biosynthesis cellular trafficking and secretion, or have a role to play in peptide stability.
The discussion above is for light chain glycosylation. It has long been known that the heavy chains of Igs are glycosylated (Alavi and Axford 1995). In each heavy chain, there is one glycosylation sequon for N-glycosylation (see Glycan-to-Protein Linkages) and this is always glycosylated, but with different glycoforms (Routier et al. 1998). In rheumatoid arthritis, there is a particular pattern of chains lacking terminal sialic acid and subterminal Gal called Gal0 and it has been postulated that the exposed GlcNAc here could have a natural GlcNAc binding ligand. It has also been suggested that the differences in glycosylation in patients with rheumatoid arthritis are responsible for changes in the protein structure and/or the resulting glycoprotein may have a different overall 3D structure with the glycans on each heavy chain interacting across the space between them.
There are several different modes of glycosylation/glycation which are implicated in pathologies associated with protein misfolding.
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