Editorial

2 pages, 182 KiB

Open AccessEditorial

Glycosylation—The Most Diverse Post-Translational Modification

by Erika Staudacher, Els J. M. Van Damme and Guy Smagghe

Biomolecules 2022, 12(9), 1313; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12091313 - 17 Sep 2022

Cited by 2 | Viewed by 1820

This article is part of the Special Issue Glycosylation—The Most Diverse Post-Translational Modification [...] Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

Research

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12 pages, 2764 KiB

Open AccessEditor’s ChoiceArticle

O-methylated N-glycans Distinguish Mosses from Vascular Plants

by David Stenitzer, Réka Mócsai, Harald Zechmeister, Ralf Reski, Eva L. Decker and Friedrich Altmann

Biomolecules 2022, 12(1), 136; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12010136 - 15 Jan 2022

Cited by 6 | Viewed by 2277

Abstract

In the animal kingdom, a stunning variety of N-glycan structures have emerged with phylogenetic specificities of various kinds. In the plant kingdom, however, N-glycosylation appears to be strictly conservative and uniform. From mosses to all kinds of gymno- and angiosperms, land plants mainly [...] Read more.

In the animal kingdom, a stunning variety of N-glycan structures have emerged with phylogenetic specificities of various kinds. In the plant kingdom, however, N-glycosylation appears to be strictly conservative and uniform. From mosses to all kinds of gymno- and angiosperms, land plants mainly express structures with the common pentasaccharide core substituted with xylose, core α1,3-fucose, maybe terminal GlcNAc residues and Lewis A determinants. In contrast, green algae biosynthesise unique and unusual N-glycan structures with uncommon monosaccharides, a plethora of different structures and various kinds of O-methylation. Mosses, a group of plants that are separated by at least 400 million years of evolution from vascular plants, have hitherto been seen as harbouring an N-glycosylation machinery identical to that of vascular plants. To challenge this view, we analysed the N-glycomes of several moss species using MALDI-TOF/TOF, PGC-MS/MS and GC-MS. While all species contained the plant-typical heptasaccharide with no, one or two terminal GlcNAc residues (MMXF, MGnXF and GnGnXF, respectively), many species exhibited MS signals with 14.02 Da increments as characteristic for O-methylation. Throughout all analysed moss N-glycans, the level of methylation differed strongly even within the same family. In some species, methylated glycans dominated, while others had no methylation at all. GC-MS revealed the main glycan from Funaria hygrometrica to contain 2,6-O-methylated terminal mannose. Some mosses additionally presented very large, likewise methylated complex-type N-glycans. This first finding of the methylation of N-glycans in land plants mirrors the presumable phylogenetic relation of mosses to green algae, where the O-methylation of mannose and many other monosaccharides is a common trait. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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20 pages, 4689 KiB

Open AccessArticle

Towards Mapping of the Human Brain N-Glycome with Standardized Graphitic Carbon Chromatography

by Johannes Helm, Lena Hirtler and Friedrich Altmann

Biomolecules 2022, 12(1), 85; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12010085 - 06 Jan 2022

Cited by 8 | Viewed by 2824

Abstract

The brain N-glycome is known to be crucial for many biological functions, including its involvement in neuronal diseases. Although large structural studies of brain N-glycans were recently carried out, a comprehensive isomer-specific structural analysis has still not been achieved, as indicated by the [...] Read more.

The brain N-glycome is known to be crucial for many biological functions, including its involvement in neuronal diseases. Although large structural studies of brain N-glycans were recently carried out, a comprehensive isomer-specific structural analysis has still not been achieved, as indicated by the recent discovery of novel structures with galactosylated bisecting GlcNAc. Here, we present a detailed, isomer-specific analysis of the human brain N-glycome based on standardized porous graphitic carbon (PGC)-LC-MS/MS. To achieve this goal, we biosynthesized glycans with substitutions typically occurring in the brain N-glycome and acquired their normalized retention times. Comparison of these values with the standardized retention times of neutral and desialylated N-glycan fractions of the human brain led to unambiguous isomer specific assignment of most major peaks. Profound differences in the glycan structures between naturally neutral and desialylated glycans were found. The neutral and sialylated N-glycans derive from diverging biosynthetic pathways and are biosynthetically finished end products, rather than just partially processed intermediates. The focus on structural glycomics defined the structure of human brain N-glycans, amongst these are HNK-1 containing glycans, a bisecting sialyl-lactose and structures with fucose and N-acetylgalactosamine on the same arm, the so-called LDNF epitope often associated with parasitic worms. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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28 pages, 6362 KiB

Open AccessEditor’s ChoiceArticle

A Combination of Structural, Genetic, Phenotypic and Enzymatic Analyses Reveals the Importance of a Predicted Fucosyltransferase to Protein O-Glycosylation in the Bacteroidetes

by Markus B. Tomek, Bettina Janesch, Matthias L. Braun, Manfred Taschner, Rudolf Figl, Clemens Grünwald-Gruber, Michael J. Coyne, Markus Blaukopf, Friedrich Altmann, Paul Kosma, Hanspeter Kählig, Laurie E. Comstock and Christina Schäffer

Biomolecules 2021, 11(12), 1795; https://0-doi-org.brum.beds.ac.uk/10.3390/biom11121795 - 30 Nov 2021

Cited by 6 | Viewed by 2563

Abstract

Diverse members of the Bacteroidetes phylum have general protein O-glycosylation systems that are essential for processes such as host colonization and pathogenesis. Here, we analyzed the function of a putative fucosyltransferase (FucT) family that is widely encoded in Bacteroidetes protein O-glycosylation [...] Read more.

Diverse members of the Bacteroidetes phylum have general protein O-glycosylation systems that are essential for processes such as host colonization and pathogenesis. Here, we analyzed the function of a putative fucosyltransferase (FucT) family that is widely encoded in Bacteroidetes protein O-glycosylation genetic loci. We studied the FucT orthologs of three Bacteroidetes species—Tannerella forsythia, Bacteroides fragilis, and Pedobacter heparinus. To identify the linkage created by the FucT of B. fragilis, we elucidated the full structure of its nine-sugar O-glycan and found that l-fucose is linked β1,4 to glucose. Of the two fucose residues in the T. forsythia O-glycan, the fucose linked to the reducing-end galactose was shown by mutational analysis to be l-fucose. Despite the transfer of l-fucose to distinct hexose sugars in the B. fragilis and T. forsythia O-glycans, the FucT orthologs from B. fragilis, T. forsythia, and P. heparinus each cross-complement the B. fragilis ΔBF4306 and T. forsythia ΔTanf_01305 FucT mutants. In vitro enzymatic analyses showed relaxed acceptor specificity of the three enzymes, transferring l-fucose to various pNP-α-hexoses. Further, glycan structural analysis together with fucosidase assays indicated that the T. forsythia FucT links l-fucose α1,6 to galactose. Given the biological importance of fucosylated carbohydrates, these FucTs are promising candidates for synthetic glycobiology. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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15 pages, 2391 KiB

Open AccessEditor’s ChoiceArticle

Assaying Paenibacillus alvei CsaB-Catalysed Ketalpyruvyltransfer to Saccharides by Measurement of Phosphate Release

by Fiona F. Hager-Mair, Cordula Stefanović, Charlie Lim, Katharina Webhofer, Simon Krauter, Markus Blaukopf, Roland Ludwig, Paul Kosma and Christina Schäffer

Biomolecules 2021, 11(11), 1732; https://0-doi-org.brum.beds.ac.uk/10.3390/biom11111732 - 20 Nov 2021

Cited by 2 | Viewed by 2197

Abstract

Ketalpyruvyltransferases belong to a widespread but little investigated class of enzymes, which utilise phosphoenolpyruvate (PEP) for the pyruvylation of saccharides. Pyruvylated saccharides play pivotal biological roles, ranging from protein binding to virulence. Limiting factors for the characterisation of ketalpyruvyltransferases are the availability of [...] Read more.

Ketalpyruvyltransferases belong to a widespread but little investigated class of enzymes, which utilise phosphoenolpyruvate (PEP) for the pyruvylation of saccharides. Pyruvylated saccharides play pivotal biological roles, ranging from protein binding to virulence. Limiting factors for the characterisation of ketalpyruvyltransferases are the availability of cognate acceptor substrates and a straightforward enzyme assay. We report on a fast ketalpyruvyltransferase assay based on the colorimetric detection of phosphate released during pyruvyltransfer from PEP onto the acceptor via complexation with Malachite Green and molybdate. To optimise the assay for the model 4,6-ketalpyruvyl::ManNAc-transferase CsaB from Paenibacillus alvei, a β-d-ManNAc-α-d-GlcNAc-diphosphoryl-11-phenoxyundecyl acceptor mimicking an intermediate of the bacterium’s cell wall glycopolymer biosynthesis pathway, upon which CsaB is naturally active, was produced chemo-enzymatically and used together with recombinant CsaB. Optimal assay conditions were 5 min reaction time at 37 °C and pH 7.5, followed by colour development for 1 h at 37 °C and measurement of absorbance at 620 nm. The structure of the generated pyruvylated product was confirmed by NMR spectroscopy. Using the established assay, the first kinetic constants of a 4,6-ketalpyuvyl::ManNAc-transferase could be determined; upon variation of the acceptor and PEP concentrations, a K_{M, PEP} of 19.50 ± 3.50 µM and k_{cat, PEP} of 0.21 ± 0.01 s⁻¹ as well as a K_{M, Acceptor} of 258 ± 38 µM and a k_{cat, Acceptor} of 0.15 ± 0.01 s⁻¹ were revealed. P. alvei CsaB was inactive on synthetic pNP-β-d-ManNAc and β-d-ManNAc-β-d-GlcNAc-1-OMe, supporting the necessity of a complex acceptor substrate. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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Review

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20 pages, 2159 KiB

Open AccessReview

Glycotherapy: A New Paradigm in Breast Cancer Research

by Dipak K. Banerjee, Arelis Seijo Lebrón and Krishna Baksi

Biomolecules 2022, 12(4), 487; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12040487 - 23 Mar 2022

Cited by 5 | Viewed by 3275

Abstract

Breast cancer is an ancient disease recognized first by the Egyptians as early as 1600 BC. The first cancer-causing gene in a chicken tumor virus was found in 1970. The United States signed the National Cancer Act in 1971, authorizing federal funding for [...] Read more.

Breast cancer is an ancient disease recognized first by the Egyptians as early as 1600 BC. The first cancer-causing gene in a chicken tumor virus was found in 1970. The United States signed the National Cancer Act in 1971, authorizing federal funding for cancer research. Irrespective of multi-disciplinary approaches, diverting a great deal of public and private resources, breast cancer remains at the forefront of human diseases, affecting as many as one in eight women during their lifetime. Because of overarching challenges and changes in the breast cancer landscape, five-year disease-free survival is no longer considered adequate. The absence of a cure, and the presence of drug resistance, severe side effects, and destruction of the patient’s quality of life, as well as the fact that therapy is often expensive, making it unaffordable to many, have created anxiety among patients, families, and friends. One of the reasons for the failure of cancer therapeutics is that the approaches do not consider cancer holistically. Characteristically, all breast cancer cells and their microenvironmental capillary endothelial cells express asparagine-linked (N-linked) glycoproteins with diverse structures. We tested a small biological molecule, Tunicamycin, that blocks a specific step of the protein N-glycosylation pathway in the endoplasmic reticulum (ER), i.e., the catalytic activity of N-acetylglusosaminyl 1-phosphate transferase (GPT). The outcome was overwhelmingly exciting. Tunicamycin quantitatively inhibits angiogenesis in vitro and in vivo, and inhibits the breast tumor progression of multiple subtypes in pre-clinical mouse models with “zero” toxicity. Mechanistic details support ER stress-induced unfolded protein response (upr) signaling as the cause for the apoptotic death of both cancer and the microvascular endothelial cells. Additionally, it interferes with Wnt signaling. We therefore conclude that Tunicamycin can be expected to supersede the current therapeutics to become a glycotherapy for treating breast cancer of all subtypes. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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10 pages, 858 KiB

Open AccessReview

Biosynthesis and Biological Significances of LacdiNAc Group on N- and O-Glycans in Human Cancer Cells

by Kiyoko Hirano and Kiyoshi Furukawa

Biomolecules 2022, 12(2), 195; https://0-doi-org.brum.beds.ac.uk/10.3390/biom12020195 - 24 Jan 2022

Cited by 12 | Viewed by 3894

Abstract

An increasing number of studies have shown that the disaccharide GalNAcβ1→4GlcNAc (LacdiNAc) group bound to N- and O-glycans in glycoproteins is expressed in a variety of mammalian cells. Biosynthesis of the LacdiNAc group was well studied, and two β4-N-acetylgalactosaminyltransferases, [...] Read more.

An increasing number of studies have shown that the disaccharide GalNAcβ1→4GlcNAc (LacdiNAc) group bound to N- and O-glycans in glycoproteins is expressed in a variety of mammalian cells. Biosynthesis of the LacdiNAc group was well studied, and two β4-N-acetylgalactosaminyltransferases, β4GalNAcT3 and β4GalNAcT4, have been shown to transfer N-acetylgalactosamine (GalNAc) to N-acetylglucosamine (GlcNAc) of N- and O-glycans in a β-1,4-linkage. The LacdiNAc group is often sialylated, sulfated, and/or fucosylated, and the LacdiNAc group, with or without these modifications, is recognized by receptors and lectins and is thus involved in the regulation of several biological phenomena, such as cell differentiation. The occurrences of the LacdiNAc group and the β4GalNAcTs appear to be tissue specific and are closely associated with the tumor progression or regression, indicating that they will be potent diagnostic markers of particular cancers, such as prostate cancer. It has been demonstrated that the expression of the LacdiNAc group on N-glycans of cell surface glycoproteins including β1-integrin is involved in the modulation of their protein functions, thus affecting cellular invasion and other malignant properties of cancer cells. The biological roles of the LacdiNAc group in cancer cells have not been fully understood. However, the re-expression of the LacdiNAc group on N-glycans, which is lost in breast cancer cells by transfection of the β4GalNAcT4 gene, brings about the partial restoration of normal properties and subsequent suppression of malignant phenotypes of the cells. Therefore, elucidation of the biological roles of the LacdiNAc group in glycoproteins will lead to the suppression of breast cancers. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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15 pages, 1000 KiB

Open AccessEditor’s ChoiceReview

Mollusc N-glycosylation: Structures, Functions and Perspectives

by Erika Staudacher

Biomolecules 2021, 11(12), 1820; https://0-doi-org.brum.beds.ac.uk/10.3390/biom11121820 - 03 Dec 2021

Cited by 10 | Viewed by 2218

Abstract

Molluscs display a sophisticated N-glycan pattern on their proteins, which is, in terms of involved structural features, even more diverse than that of vertebrates. This review summarises the current knowledge of mollusc N-glycan structures, with a focus on the functional aspects of the [...] Read more.

Molluscs display a sophisticated N-glycan pattern on their proteins, which is, in terms of involved structural features, even more diverse than that of vertebrates. This review summarises the current knowledge of mollusc N-glycan structures, with a focus on the functional aspects of the corresponding glycoproteins. Furthermore, the potential of mollusc-derived biomolecules for medical applications is addressed, emphasising the importance of mollusc research. Full article

(This article belongs to the Special Issue Glycosylation—The Most Diverse Post-Translational Modification)

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