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  • Thiomyristoyl Finally the formation of a regular network

    2021-09-14

    Finally, the formation of a regular network of fusion glycoproteins in their post-fusion state is not exclusive of vesiculovirus G. More or less regular networks have been also observed with other class III fusion glycoproteins such as pseudorabies gB [85] and several class II fusion glycoproteins [51], [86], [87]. As suggested for vesiculoviruses [38], it is possible that the glycoproteins located outside the contact zone between fusing membranes have a role, probably at a late stage of the fusion process (i.e., enlargement of the fusion pore) via the formation of more or less regular arrays.
    Final remarks and conclusion Novel structures of such intermediates are required to complete our understanding of the transition pathway for each class of viral fusion glycoproteins. Cryo-EM and cryo-ET enhanced by direct Thiomyristoyl detection devices, improved microscopes with more stable optics, and advances in image processing software [88], [89], [90], [91] should make it possible to visualize the conformation of the glycoproteins while they interact with a target membrane. Indeed, as mentioned above, some progresses have already been made for both class I [81], [82], [83], [84], [92] and class II fusion glycoproteins [67], [68], [69]; however, a higher resolution will be required to get a reliable quasi-atomic model of those intermediates.
    Acknowledgments We thank Nathan Jespersen for providing language help and careful reading of the manuscript. This work was supported by a grant from ANR (ANR CE11 MOBARHE) to Y.G.
    Introduction The glycosylation of proteins is one of the most frequent posttranslational modifications, which is not well understood on a functional level. For mammalian glycoproteins the carbohydrate part is known to affect numerous biological processes in a direct or indirect manner. Thus, the individual glycosylation pattern of a protein is of significant importance for both protein structure and function. The carbohydrate structures of individual glycoforms are of importance for the in vivo activity of recombinant therapeutic glycoproteins, for example, erythropoietin and therapeutic antibodies. Asparagine-linked oligosaccharides (N-glycans) can also confer stability to a protein by means of intramolecular protein–carbohydrate interactions. An insightful and comprehensive overview of this area is given in the review by Varki [1]. However, due to the heterogeneity of natural glycoproteins (glycoforms) in the sugar part the evaluation of the impact of individual glycans remains difficult to investigate. Pure glycoforms of glycoproteins are still not accessible via recombinant or chromatographic methods. Thus, the synthesis of N-glycoproteins with uniform posttranslational modifications using ligation methods [2, 3, 4] or enzymatic remodeling of N-glycans [5, 6] are currently the best options for accessing pure glycoforms (Figure 1). Only recently the combination of many advances in this area enabled the first total syntheses of pure N-glycoproteins []. These approaches have quickly matured to a very dynamic research field, providing natural structures as well as their analogs. The use of endo-β-N-acetylglucosaminidases (ENGases) can provide rapid access to pure glycoforms. The cleavage of the chitobiose core of N-glycans is reversible and can be used for synthesis when highly reactive oxazolines are employed as substrates. When utilizing wild type ENGases the extent of hydrolysis of the newly formed glycoform must be carefully monitored in order to optimise the yield of the enzymatic transfer reaction. Additionally there is a strong preference of each ENGase for certain types of N-glycans [8]. Thus, ENGases with mutations of the catalytic residues have been developed, which act only as a synthase and permit high conversion without hydrolysis of the final product [5,[]]. A limitation of this otherwise very promising approach was recently discovered. Stoichiometric amounts of N-glycan-oxazolines may also react non-specifically [] with amino groups [10] of the acceptor protein or peptide.