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ACS Synthetic Biology Jul 2020Protein glycosylation, the attachment of sugars to amino acid side chains, can endow proteins with a wide variety of properties of great interest to the engineering... (Review)
Review
Protein glycosylation, the attachment of sugars to amino acid side chains, can endow proteins with a wide variety of properties of great interest to the engineering biology community. However, natural glycosylation systems are limited in the diversity of glycoproteins they can synthesize, the scale at which they can be harnessed for biotechnology, and the homogeneity of glycoprotein structures they can produce. Here we provide an overview of the emerging field of synthetic glycobiology, the application of synthetic biology tools and design principles to better understand and engineer glycosylation. Specifically, we focus on how the biosynthetic and analytical tools of synthetic biology have been used to redesign glycosylation systems to obtain defined glycosylation structures on proteins for diverse applications in medicine, materials, and diagnostics. We review the key biological parts available to synthetic biologists interested in engineering glycoproteins to solve compelling problems in glycoscience, describe recent efforts to construct synthetic glycoprotein synthesis systems, and outline exemplary applications as well as new opportunities in this emerging space.
Topics: Animals; Bacteria; Biotechnology; Fungi; Glycosylation; Glycosyltransferases; Plants; Proteins; Synthetic Biology
PubMed: 32526139
DOI: 10.1021/acssynbio.0c00210 -
Theranostics 2023Cell surface glycosylation has a variety of functions, and its dysregulation in cancer contributes to impaired signaling, metastasis and the evasion of the immune... (Review)
Review
Cell surface glycosylation has a variety of functions, and its dysregulation in cancer contributes to impaired signaling, metastasis and the evasion of the immune responses. Recently, a number of glycosyltransferases that lead to altered glycosylation have been linked to reduced anti-tumor immune responses: B3GNT3, which is implicated in PD-L1 glycosylation in triple negative breast cancer, FUT8, through fucosylation of B7H3, and B3GNT2, which confers cancer resistance to T cell cytotoxicity. Given the increased appreciation of the relevance of protein glycosylation, there is a critical need for the development of methods that allow for an unbiased interrogation of cell surface glycosylation status. Here we provide an overview of the broad changes in glycosylation at the surface of cancer cell and describe selected examples of receptors with aberrant glycosylation leading to functional changes, with emphasis on immune checkpoint inhibitors, growth-promoting and growth-arresting receptors. Finally, we posit that the field of glycoproteomics has matured to an extent where large-scale profiling of intact glycopeptides from the cell surface is feasible and is poised for discovery of new actionable targets against cancer.
Topics: Humans; Glycosylation; Cell Membrane; Glycopeptides; Triple Negative Breast Neoplasms; Carrier Proteins
PubMed: 37215580
DOI: 10.7150/thno.81760 -
Gastroenterology Jan 2020Glycans are sequences of carbohydrates that are added to proteins or lipids to modulate their structure and function. Glycans modify proteins required for regulation of... (Review)
Review
Glycans are sequences of carbohydrates that are added to proteins or lipids to modulate their structure and function. Glycans modify proteins required for regulation of immune cells, and alterations have been associated with inflammatory conditions. For example, specific glycans regulate T-cell activation, structures, and functions of immunoglobulins; interactions between microbes and immune and epithelial cells; and malignant transformation in the intestine and liver. We review the effects of protein glycosylation in regulation of gastrointestinal and liver functions, and how alterations in glycosylation serve as diagnostic or prognostic factors, or as targets for therapy.
Topics: Biomarkers; Gastrointestinal Diseases; Gastrointestinal Tract; Glycomics; Glycosylation; Humans; Liver; Liver Diseases; Polysaccharides; Prognosis; Proteomics; T-Lymphocytes; Time Factors
PubMed: 31626754
DOI: 10.1053/j.gastro.2019.08.060 -
Future Microbiology Jan 2017Through advances in analytical methods to detect glycoproteins and to determine glycan structures, there have been increasing reports of protein glycosylation in... (Review)
Review
Through advances in analytical methods to detect glycoproteins and to determine glycan structures, there have been increasing reports of protein glycosylation in bacteria. In this review, we summarize the known pathways for bacterial protein glycosylation: lipid carrier-mediated 'en bloc' glycosylation; and cytoplasmic stepwise protein glycosylation. The exploitation of bacterial protein glycosylation systems, especially the 'mix and match' of three independent but similar pathways (oligosaccharyltransferase-mediated protein glycosylation, lipopolysaccharide and peptidoglycan biosynthesis) in Gram-negative bacteria for glycoengineering recombinant glycoproteins is also discussed.
Topics: Bacterial Proteins; Campylobacter jejuni; Glycosylation; Gram-Positive Bacteria; Helicobacter pylori; Hexosyltransferases; Lipopolysaccharides; Membrane Proteins; Neisseria gonorrhoeae; Neisseria meningitidis; Peptidoglycan; Polysaccharides; Protein Engineering; Recombinant Proteins
PubMed: 27689684
DOI: 10.2217/fmb-2016-0166 -
Postepy Biochemii Sep 2023Glycosylated proteins play a key role in the various stages of bacterial and viral invasions. Glycosylation is a common process across all domains of life. Initially,...
Glycosylated proteins play a key role in the various stages of bacterial and viral invasions. Glycosylation is a common process across all domains of life. Initially, this process was attributed only to eukaryotic organisms, in which the synthesis takes place in the rough endoplasmic reticulum and the Golgi apparatus. Over time, it has been shown that many bacteria and viruses express N-glycans and O-glycans on their surface. Prokaryotes are able to synthesize glycans, while virions take over the host's cellular machinery to produce glycans. Pathogens use glycoproteins to regulate adhesion to infected cells (Ebola virus), protect receptor-binding epitopes (HIV) and evade the immune system detection by molecular mimicry (Helicobacter pylori, Haemophilus influenzae). Successful infection also depends on the host surface glycans, mainly in determining the tissue tropism of viruses (Influenza A viruses) and the sliding motility of bacteria (Mycoplasma sp.). Modification of glycan structures, important at various levels of the infectious cycle, creates new therapeutic possibilities that gives a chance to limit the spread of infectious diseases.
Topics: Humans; Glycosylation; Viruses; Polysaccharides; Virus Diseases; Bacteria
PubMed: 38019747
DOI: 10.18388/pb.2021_488 -
Current Opinion in Structural Biology Jun 2022Biosynthetic enzymes in the secretory pathway create distributions of glycans at each glycosite that elaborate the biophysical properties and biological functions of... (Review)
Review
Biosynthetic enzymes in the secretory pathway create distributions of glycans at each glycosite that elaborate the biophysical properties and biological functions of glycoproteins. Because the biosynthetic glycosylation reactions do not go to completion, each protein glycosite is heterogeneous with respect to glycosylation. This heterogeneity means that it is not sufficient to measure protein abundance in omics experiments. Rather, it is necessary to sample the distribution of glycosylation at each glycosite to quantify the changes that occur during biological processes. On the one hand, the use of data-dependent acquisition methods to sample glycopeptides is limited by the instrument duty cycle and the missing value problem. On the other, stepped window data-independent acquisition samples all precursors, but ion abundances are limited by duty cycle. Therefore, the ability to quantify accurately the flux in glycoprotein glycosylation that occurs during biological processes requires the exploitation of emerging mass spectrometry technologies capable of deep, comprehensive sampling and selective high confidence assignment of the complex glycopeptide mixtures. This review summarizes recent technical advances and mass spectral glycoproteomics analysis strategies and how these developments impact our ability to quantify the changes in glycosylation that occur during biological processes. We highlight specific improvements to glycopeptide characterization through activated electron dissociation, ion mobility trends and instrumentation, and efficient algorithmic approaches for glycopeptide assignment. We also discuss the emerging need for unified standards to enable interlaboratory collaborations and effective monitoring of structural changes in glycoproteins.
Topics: Glycopeptides; Glycoproteins; Glycosylation; Mass Spectrometry; Polysaccharides
PubMed: 35452871
DOI: 10.1016/j.sbi.2022.102371 -
Molecules (Basel, Switzerland) Nov 2018-Glycosylation in general has impact on a diversity of biological processes covering cellular aspects (targeted transport of glycoproteins), molecular aspects (protein...
-Glycosylation in general has impact on a diversity of biological processes covering cellular aspects (targeted transport of glycoproteins), molecular aspects (protein conformation, resistance to proteolysis), and aspects involved in cellular communication (cell-cell and cell-matrix interaction). [...].
Topics: Animals; Glycoproteins; Glycosylation; Humans; Protein Processing, Post-Translational
PubMed: 30477085
DOI: 10.3390/molecules23123063 -
Mass Spectrometry-Based Chemical and Enzymatic Methods for Global Analysis of Protein Glycosylation.Accounts of Chemical Research Aug 2018Glycosylation is one of the most common protein modifications, and it is essential for mammalian cell survival. It often determines protein folding and trafficking, and... (Review)
Review
Glycosylation is one of the most common protein modifications, and it is essential for mammalian cell survival. It often determines protein folding and trafficking, and regulates nearly every extracellular activity, including cell-cell communication and cell-matrix interactions. Aberrant protein glycosylation events are hallmarks of human diseases such as cancer and infectious diseases. Therefore, glycoproteins can serve as effective biomarkers for disease detection and targets for drug and vaccine development. Despite the importance of glycoproteins, global analysis of protein glycosylation (either glycoproteins or glycans) in complex biological samples has been a daunting task, and here we mainly focus on glycoprotein analysis using mass spectrometry (MS)-based bottom-up proteomics. Although the emergence of MS-based proteomics has provided a great opportunity to analyze glycoproteins globally, the low abundance of many glycoproteins and the heterogeneity of glycans dramatically increase the technical difficulties. In order to overcome these obstacles, considerable progress has been made in recent years, which has contributed to comprehensive analysis of glycoproteins. In our lab, we developed effective MS-based chemical and enzymatic methods to (1) globally analyze glycoproteins in complex biological samples, (2) target glycoproteins specifically on the surface of human cells, (3) systematically quantify glycoprotein and surface glycoprotein dynamics (the abundance changes of glycoproteins as a function of time), and (4) selectively characterize glycoproteins with a particular and important glycan. In this Account, we first briefly describe the glycopeptide/protein enrichment methods in the literature and then discuss the developments of boronic acid-based methods to enrich glycopeptides for large-scale analysis of protein glycosylation. Boronic acids can form reversible covalent interactions with sugars, but the low binding affinity of normal boronic acid-based methods prevents us from capturing glycoproteins with low abundance, which often contain more valuable information. We enhanced the boronic acid-glycan interactions by using a boronic acid derivative (benzoboroxole) and conjugating it onto a dendrimer to allow synergistic interactions between the boronic acid derivative and sugars. The new method is capable of globally analyzing protein glycosylation with site and glycan structure information, especially for those with low abundance. In the next part, we discuss the combination of metabolic labeling, click chemistry and enzymatic reactions, and MS-based proteomics as a very powerful approach for surface glycoproteome analysis in human cells. The methods enable us to specifically identify surface glycoproteins and to quantify their abundance changes and dynamics together with quantitative proteomics. The last section of this Account focuses on chemical and enzymatic methods to study glycoproteins containing a particular and important glycan (the Tn antigen, i.e., O-GalNAc). Although not comprehensive, this Account provides an overview of chemical and enzymatic methods to characterize protein glycosylation in combination with MS-based proteomics. These methods will have extensive applications in the fields of biology and biomedicine, which will lead to a better understanding of glycoprotein functions and the molecular mechanisms of diseases. Eventually, glycoproteins will be identified as effective biomarkers for disease detection and drug targets for disease treatment.
Topics: Antigens, Tumor-Associated, Carbohydrate; Boronic Acids; Click Chemistry; Glycosylation; Humans; Mass Spectrometry; Membrane Glycoproteins; Polysaccharides; Protein Processing, Post-Translational; Proteome; Proteomics
PubMed: 30011186
DOI: 10.1021/acs.accounts.8b00200 -
Cells May 2021Evading host immune surveillance is one of the hallmarks of cancer. Immune checkpoint therapy, which aims to eliminate cancer progression by reprogramming the antitumor... (Review)
Review
Evading host immune surveillance is one of the hallmarks of cancer. Immune checkpoint therapy, which aims to eliminate cancer progression by reprogramming the antitumor immune response, currently occupies a solid position in the rapidly expanding arsenal of cancer therapy. As most immune checkpoints are membrane glycoproteins, mounting attention is drawn to asking how protein glycosylation affects immune function. The answers to this fundamental question will stimulate the rational development of future cancer diagnostics and therapeutic strategies.
Topics: Animals; Glycosylation; Humans; Immune Checkpoint Inhibitors; Neoplasms; Protein Processing, Post-Translational; Receptors, Immunologic
PubMed: 34064396
DOI: 10.3390/cells10051100 -
Current Opinion in Structural Biology Apr 2023Glycosyltransferases of the C superfamily (GT-Cs) are enzymes found in all domains of life. They catalyse the stepwise synthesis of oligosaccharides or the transfer of... (Review)
Review
Glycosyltransferases of the C superfamily (GT-Cs) are enzymes found in all domains of life. They catalyse the stepwise synthesis of oligosaccharides or the transfer of assembled glycans from lipid-linked donor substrates to acceptor proteins. The processes mediated by GT-Cs are required for C-, N- and O-linked glycosylation, all of which are essential post-translational modifications in higher-order eukaryotes. Until recently, GT-Cs were thought to share a conserved structural module of 7 transmembrane helices; however, recently determined GT-C structures revealed novel folds. Here we analyse the growing diversity of GT-C folds and discuss the emergence of two subclasses, termed GT-C and GT-C. Further substrate-bound structures are needed to facilitate a molecular understanding of glycan recognition and catalysis in these two subclasses.
Topics: Glycosyltransferases; Glycosylation; Polysaccharides; Oligosaccharides; Protein Structure, Secondary
PubMed: 36827761
DOI: 10.1016/j.sbi.2023.102547