Publications by authors named "Alba Silipo"

135 Publications

Recent advances on smart glycoconjugate vaccines in infections and cancer.

FEBS J 2021 May 2. Epub 2021 May 2.

Department of Molecular Cell Biology and Immunology, Cancer Center Amsterdam, Amsterdam Infection and Immunity Institute, Amsterdam UMC, Vrije Universiteit Amsterdam, The Netherlands.

Vaccination is one of the greatest achievements in biomedical research preventing death and morbidity in many infectious diseases through the induction of pathogen-specific humoral and cellular immune responses. Currently, no effective vaccines are available for pathogens with a highly variable antigenic load, such as the human immunodeficiency virus or to induce cellular T-cell immunity in the fight against cancer. The recent SARS-CoV-2 outbreak has reinforced the relevance of designing smart therapeutic vaccine modalities to ensure public health. Indeed, academic and private companies have ongoing joint efforts to develop novel vaccine prototypes for this virus. Many pathogens are covered by a dense glycan-coat, which form an attractive target for vaccine development. Moreover, many tumor types are characterized by altered glycosylation profiles that are known as "tumor-associated carbohydrate antigens". Unfortunately, glycans do not provoke a vigorous immune response and generally serve as T-cell-independent antigens, not eliciting protective immunoglobulin G responses nor inducing immunological memory. A close and continuous crosstalk between glycochemists and glycoimmunologists is essential for the successful development of efficient immune modulators. It is clear that this is a key point for the discovery of novel approaches, which could significantly improve our understanding of the immune system. In this review, we discuss the latest advancements in development of vaccines against glycan epitopes to gain selective immune responses and to provide an overview on the role of different immunogenic constructs in improving glycovaccine efficacy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/febs.15909DOI Listing
May 2021

Investigation of protein-ligand complexes by ligand-based NMR methods.

Carbohydr Res 2021 May 9;503:108313. Epub 2021 Apr 9.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Via Cintia 4, I-80126, Napoli, Italy; CNR, Institute for Polymers, Composites and Biomaterials, IPCB ss, Catania, Italy. Electronic address:

Molecular recognition is at the base of all biological events and its knowledge at atomic level is pivotal in the development of new drug design approaches. NMR spectroscopy is one of the most widely used technique to detect and characterize transient ligand-receptor interactions in solution. In particular, ligand-based NMR approaches, including NOE-based NMR techniques, diffusion experiments and relaxation methods, are excellent tools to investigate how ligands interact with their receptors. Here we describe the key structural information that can be achieved on binding processes thanks to the combined used of advanced NMR and computational methods. Saturation Transfer Difference NMR (STD-NMR), WaterLOGSY, diffusion- and relaxation-based experiments, together with tr-NOE techniques allow, indeed, to investigate the ligand behavior when bound to a receptor, determining, among others, the epitope map of the ligand and its bioactive conformation. The combination of these NMR techniques with computational methods, including docking, molecular dynamics and CORCEMA-ST analysis, permits to define and validate an accurate 3D model of protein-ligand complexes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.carres.2021.108313DOI Listing
May 2021

Covalently bonded hopanoid-Lipid A from Bradyrhizobium: The role of unusual molecular structure and calcium ions in regulating the lipid bilayers organization.

J Colloid Interface Sci 2021 Jul 18;594:891-901. Epub 2021 Mar 18.

CSGI, Center for Colloid and Surface Science, via della Lastruccia 3, Sesto Fiorentino (FI), Italy; Department of Chemical Sciences, University of Naples Federico II, via Cintia 4, 80126 Naples, Italy. Electronic address:

Lipopolysaccharides (LPS) are complex amphiphilic macromolecules forming the external leaflet of the outer membrane of Gram-negative bacteria. The LPS glycolipid portion, named Lipid A, is characterized by a disaccharide backbone carrying multiple acyl chains. Some Lipid A bear very-long-chain-fatty-acids (VLCFA), biosynthesized to span the entire lipid membrane profile. The synbiontic Bradyrhizobium BTAi1 strain carries an unique Lipid A specie, named HoLA, in which VLCFA terminus is covalently-bonded to hopanoid, a triterpenoid displaying structural similarity with eukaryotic sterols. Here, we investigate the role of HoLA in regulating self-assembly, microstructure and thermotropicity of lipid membranes composed by 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-1'-rac-glycerol. DLS and SANS indicated the formation of multilamellar aggregates whose size increases when the hopanoid is present in the bilayer. EPR and DSC showed that HoLA induced a more rigid and ordered organization among the lipids in the bilayer, evocative of a mechanical strengthening. Notably, the presence of calcium ions promote an increase of the bilayer thickness and vesicles size, leading to low curvature aggregates. These results highlight the key role of the hopanoid covalently-linked to VLCFA in defining the physico-chemical properties of bacterial envelope, thus offering a robust scientific basis for the interpretation of the biological activity of the considered strain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jcis.2021.03.072DOI Listing
July 2021

Emerging glyco-based strategies to steer immune responses.

FEBS J 2021 Mar 22. Epub 2021 Mar 22.

Department of Molecular Cell Biology and Immunology, Cancer Center Amsterdam, Amsterdam Infection and Immunity Institute, Amsterdam UMC, Vrije Universiteit Amsterdam, Netherlands.

Glycan structures are common posttranslational modifications of proteins, which serve multiple important structural roles (for instance in protein folding), but also are crucial participants in cell-cell communications and in the regulation of immune responses. Through the interaction with glycan-binding receptors, glycans are able to affect the activation status of antigen-presenting cells, leading either to induction of pro-inflammatory responses or to suppression of immunity and instigation of immune tolerance. This unique feature of glycans has attracted the interest and spurred collaborations of glyco-chemists and glyco-immunologists to develop glycan-based tools as potential therapeutic approaches in the fight against diseases such as cancer and autoimmune conditions. In this review, we highlight emerging advances in this field, and in particular, we discuss on how glycan-modified conjugates or glycoengineered cells can be employed as targeting devices to direct tumor antigens to lectin receptors on antigen-presenting cells, like dendritic cells. In addition, we address how glycan-based nanoparticles can act as delivery platforms to enhance immune responses. Finally, we discuss some of the latest developments in glycan-based therapies, including chimeric antigen receptor (CAR)-T cells to achieve targeting of tumor-associated glycan-specific epitopes, as well as the use of glycan moieties to suppress ongoing immune responses, especially in the context of autoimmunity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/febs.15830DOI Listing
March 2021

Understanding the Antibacterial Resistance: Computational Explorations in Bacterial Membranes.

ACS Omega 2021 Mar 26;6(9):6041-6054. Epub 2021 Feb 26.

Department of Structural and Chemical Biology, Centro de Investigaciones Biológicas Research Margarita Salas, CIB-CSIC, C/Ramiro de Maeztu, 9, 28040 Madrid, Spain.

Antimicrobial resistance (AMR) represents a major threat to global public health in the 21st century, dramatically increasing the pandemic expectations in the coming years. The ongoing need to develop new antimicrobial treatments that are effective against multi-drug-resistant pathogens has led the research community to investigate innovative strategies to tackle AMR. The bacterial cell envelope has been identified as one of the key molecular players responsible for antibiotic resistance, attracting considerable interest as a potential target for novel antimicrobials effective against AMR, to be used alone or in combination with other drugs. However, the multicomponent complexity of bacterial membranes provides a heterogeneous morphology, which is typically difficult to study at the molecular level by experimental techniques, in spite of the significant development of fast and efficient experimental protocols. In recent years, computational modeling, in particular, molecular dynamics simulations, has proven to be an effective tool to reveal key aspects in the architecture and membrane organization of bacterial cell walls. Here, after a general overview about bacterial membranes, AMR mechanisms, and experimental approaches to study AMR, we review the state-of-the-art computational approaches to investigate bacterial AMR envelopes, including their limitations and challenges ahead. Representative examples illustrate how these techniques improve our understanding of bacterial membrane resistance mechanisms, hopefully leading to the development of novel antimicrobial drugs escaping from bacterial resistance strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acsomega.0c05590DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7948216PMC
March 2021

Analysis of Synthetic Monodisperse Polysaccharides by Wide Mass Range Ultrahigh-Resolution MALDI Mass Spectrometry.

Anal Chem 2021 03 5;93(10):4666-4675. Epub 2021 Mar 5.

Center for Proteomics and Metabolomics, Leiden University Medical Center, Leiden 2333 ZA, The Netherlands.

Carbohydrates, such as oligo- and polysaccharides, are highly abundant biopolymers that are involved in numerous processes. The study of their structure and functions is commonly based on a material that is isolated from complex natural sources. However, a more precise analysis requires pure compounds with well-defined structures that can be obtained from chemical or enzymatic syntheses. Novel synthetic strategies have increased the accessibility of larger monodisperse polysaccharides, posing a challenge to the analytical methods used for their molecular characterization. Here, we present wide mass range ultrahigh-resolution matrix-assisted laser desorption/ionization (MALDI) Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS) as a powerful platform for the analysis of synthetic oligo- and polysaccharides. Synthetic carbohydrates 16-, 64-, 100-, and 151-mers were mass analyzed and characterized by MALDI in-source decay FT-ICR MS. Detection of fragment ions generated from glycosidic bond cleavage (or cross-ring cleavage) provided information of the monosaccharide content and the linkage type, allowing for the corroboration of the carbohydrate compositions and structures.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.analchem.1c00239DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8034773PMC
March 2021

Lipopolysaccharide from Gut-Associated Lymphoid-Tissue-Resident Alcaligenes faecalis: Complete Structure Determination and Chemical Synthesis of Its Lipid A.

Angew Chem Int Ed Engl 2021 04 22;60(18):10023-10031. Epub 2021 Mar 22.

Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka, 560-0043, Japan.

Alcaligenes faecalis is the predominant Gram-negative bacterium inhabiting gut-associated lymphoid tissues, Peyer's patches. We previously reported that an A. faecalis lipopolysaccharide (LPS) acted as a weak agonist for Toll-like receptor 4 (TLR4)/myeloid differentiation factor-2 (MD-2) receptor as well as a potent inducer of IgA without excessive inflammation, thus suggesting that A. faecalis LPS might be used as a safe adjuvant. In this study, we characterized the structure of both the lipooligosaccharide (LOS) and LPS from A. faecalis. We synthesized three lipid A molecules with different degrees of acylation by an efficient route involving the simultaneous introduction of 1- and 4'-phosphates. Hexaacylated A. faecalis lipid A showed moderate agonistic activity towards TLR4-mediated signaling and the ability to elicit a discrete interleukin-6 release in human cell lines and mice. It was thus found to be the active principle of the LOS/LPS and a promising vaccine adjuvant candidate.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/anie.202012374DOI Listing
April 2021

Behavior of glycolylated sialoglycans in the binding pockets of murine and human CD22.

iScience 2021 Jan 29;24(1):101998. Epub 2020 Dec 29.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Via Cintia 4, 80126 Napoli, Italy.

Siglecs (sialic acid binding immunoglobulin (Ig)-like lectins) constitute a group of 15 human and 9 murine cell-surface transmembrane receptors belonging to the I-type lectin family, mostly expressed on innate immune cells and characterized by broadly similar structural features. Here, the prominent inhibitory CD22 (Siglec-2), well known in maintaining tolerance and preventing autoimmune responses on B cells, is studied in its human and murine forms in complex with sialoglycans. In detail, the role of the -glycolyl neuraminic acid (Neu5Gc) moiety in the interaction with both orthologues was explored. The analysis of the binding mode was carried out by the combination of NMR spectroscopy, computational approaches, and CORCEMA-ST calculations. Our findings provide a first model of Neu5Gc recognition by h-CD22 and show a comparable molecular recognition profile by h- and m-CD22. These data open the way to innovative diagnostic and/or therapeutic methodologies to be used in the modulation of the immune responses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.isci.2020.101998DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7811138PMC
January 2021

Solving the structural puzzle of bacterial glycome.

Curr Opin Struct Biol 2021 Jan 9;68:74-83. Epub 2021 Jan 9.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Via Cintia 4, I-80126 Napoli, Italy. Electronic address:

The analysis of the bacterial glycome (glycomics) is among the complex 'omics' analysis owing to the inherent difficulties in structural and functional characterization of glycans. The complexity and variability of bacterial glycans, spanning from simple carbohydrates to complex glycolipids, glycopeptides and glycoproteins, make their study a challenging research area. The last two decades have witnessed tremendous advances and development of highly sophisticated methods, in combination with optimized protocols and hyphenate techniques for the understanding of structure, conformations, dynamics and organization of microbial glycans. We here present an overview of the novel approaches that have massively improved our understanding of the carbohydrate-based world of bacteria.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.sbi.2020.12.003DOI Listing
January 2021

The structure of SeviL, a GM1b/asialo-GM1 binding R-type lectin from the mussel Mytilisepta virgata.

Sci Rep 2020 12 16;10(1):22102. Epub 2020 Dec 16.

Graduate School of Medical Life Science, Yokohama City University, 1-7-29 Suehiro, Yokohama, Kanagawa, 230-0045, Japan.

SeviL is a recently isolated lectin found to bind to the linear saccharides of the ganglioside GM1b (Neu5Ac[Formula: see text](2-3)Gal[Formula: see text](1-3)GalNAc[Formula: see text](1-4)Gal[Formula: see text](1-4)Glc) and its precursor, asialo-GM1 (Gal[Formula: see text](1-3)GalNAc[Formula: see text](1-4)Gal[Formula: see text](1-4)Glc). The crystal structures of recombinant SeviL have been determined in the presence and absence of ligand. The protein belongs to the [Formula: see text]-trefoil family, but shows only weak sequence similarity to known structures. SeviL forms a dimer in solution, with one binding site per subunit, close to the subunit interface. Molecular details of glycan recognition by SeviL in solution were analysed by ligand- and protein-based NMR techniques as well as ligand binding assays. SeviL shows no interaction with GM1 due to steric hindrance with the sialic acid branch that is absent from GM1b. This unusual specificity makes SeviL of great interest for the detection and control of certain cancer cells, and cells of the immune system, that display asialo-GM1.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-78926-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7744527PMC
December 2020

The Structure of the Lipid A of Gram-Negative Cold-Adapted Bacteria Isolated from Antarctic Environments.

Mar Drugs 2020 Nov 26;18(12). Epub 2020 Nov 26.

Department of Chemical Sciences, University of Napoli Federico II, Complesso Universitario Monte S. Angelo, Via Cintia 4, I-80126 Napoli, Italy.

Gram-negative Antarctic bacteria adopt survival strategies to live and proliferate in an extremely cold environment. Unusual chemical modifications of the lipopolysaccharide (LPS) and the main component of their outer membrane are among the tricks adopted to allow the maintenance of an optimum membrane fluidity even at particularly low temperatures. In particular, the LPS' glycolipid moiety, the lipid A, typically undergoes several structural modifications comprising desaturation of the acyl chains, reduction in their length and increase in their branching. The investigation of the structure of the lipid A from cold-adapted bacteria is, therefore, crucial to understand the mechanisms underlying the cold adaptation phenomenon. Here we describe the structural elucidation of the highly heterogenous lipid A from three psychrophiles isolated from Terra Nova Bay, Antarctica. All the lipid A structures have been determined by merging data that was attained from the compositional analysis with information from a matrix-assisted laser desorption ionization (MALDI) time of flight (TOF) mass spectrometry (MS) and MS investigation. As lipid A is also involved in a structure-dependent elicitation of innate immune response in mammals, the structural characterization of lipid A from such extremophile bacteria is also of great interest from the perspective of drug synthesis and development inspired by natural sources.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/md18120592DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7759928PMC
November 2020

A chronic strain of the cystic fibrosis pathogen Pandoraea pulmonicola expresses a heterogenous hypo-acylated lipid A.

Glycoconj J 2021 Apr 13;38(2):135-144. Epub 2020 Oct 13.

Department of Chemical Sciences, University of Naples Federico II, via Cinthia 4, Naples, 80126, Italy.

Pandoraea sp. is an emerging Gram-negative pathogen in cystic fibrosis causing severe and persistent inflammation and damage of the lungs. The molecular mechanisms underlying the high pathogenicity of Pandoraea species are still largely unknown. As Gram-negatives, Pandoraea sp. express lipopolysaccharides (LPS) whose recognition by the host immune system triggers an inflammatory response aimed at the bacterial eradication from the infected tissues. The degree of the inflammatory response strongly relies on the fine structure of the LPS and, in particular, of its glycolipid moiety, i.e. the lipid A. Here we report the structure of the lipid A isolated from the LPS of a chronic strain of P. pulmonicola (RL 8228), one of the most virulent identified so far among the Pandoraea species. Our data demonstrated that the examined chronic strain produces a smooth-type LPS with a complex mixture of hypoacylated lipid A species displaying, among other uncommon characteristics, the 2-hydroxylation of some of the acyl chains and the substitution by an additional glucosamine on one or both the phosphate groups.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10719-020-09954-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8052242PMC
April 2021

Pairing LPS Structure with Its Immunomodulatory Effects on Human Cellular Models.

ACS Cent Sci 2020 Sep 30;6(9):1602-1616. Epub 2020 Jul 30.

Department of Chemical Sciences, University of Naples Federico II, 80126 Naples, Italy.

The gut microbiota guide the development of the host immune system by setting a systemic threshold for immune activation. Lipopolysaccharides (LPSs) from gut bacteria are able to trigger systemic and local proinflammatory and immunomodulatory responses, and this capability strongly relies on their fine structures. Up to now, only a few LPS structures from gut commensals have been elucidated; therefore, the molecular motifs that may be important for LPS-mammalian cell interactions at the gut level are still obscure. Here, we report on the full structure of the LPS isolated from one of the prominent species of the genus , . The LPS turned out to consist of a particular chemical structure based on hypoacylated and -phosphorylated lipid A and with a galactofuranose-containing core oligosaccharide and an O-antigen built up of mannose and rhamnose. The evaluation of the immunological properties of this LPS on human models revealed a very interesting capability to produce anti-inflammatory cytokines and to induce a synergistic action of MD-2/TLR4- and TLR2-mediated signaling pathways.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acscentsci.0c00791DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7517413PMC
September 2020

The Peculiar Structure of Acetobacter pasteurianus CIP103108 LPS Core Oligosaccharide.

Chembiochem 2021 Jan 29;22(1):147-150. Epub 2020 Oct 29.

Department of Chemical Sciences, University of Naples Federico II Complesso Universitario Monte Sant'Angelo, Via Cintia 4, 80126, Napoli, Italy.

Acetobacter pasteurianus, a member of the Alphaproteobacteria, is an acetic acid-producing bacterium present on sugar-rich substrates such as such as fruits, flowers and vegetables and traditionally used in the production of fermented food. The preferred living habitat associated with acid conditions makes the structure of the bacterial cell wall interesting to study, due to expected uncommon features. We have used a combination of chemical, analytical and NMR spectroscopy approaches to define the complete structure of the core oligosaccharide from A. pasteurianus CIP103108 LPS. Interestingly, the core oligosaccharide displays a high concentration of negatively charged groups, structural features that might contribute to reinforcing the bacterial membrane.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/cbic.202000597DOI Listing
January 2021

Chemical synthesis of glycans up to a 128-mer relevant to the O-antigen of Bacteroides vulgatus.

Nat Commun 2020 08 18;11(1):4142. Epub 2020 Aug 18.

State Key Laboratory of Bioorganic and Natural Products Chemistry, Center for Excellence in Molecular Synthesis, Shanghai Institute of Organic Chemistry, University of Chinese Academy of Sciences, Chinese Academy of Sciences, 345 Lingling Road, Shanghai, 200032, China.

Glycans are involved in various life processes and represent critical targets of biomedical developments. Nevertheless, the accessibility to long glycans with precise structures remains challenging. Here we report on the synthesis of glycans consisting of [→4)-α-Rha-(1 → 3)-β-Man-(1 → ] repeating unit, which are relevant to the O-antigen of Bacteroides vulgatus, a common component of gut microbiota. The optimal combination of assembly strategy, protecting group arrangement, and glycosylation reaction has enabled us to synthesize up to a 128-mer glycan. The synthetic glycans are accurately characterized by advanced NMR and MS approaches, the 3D structures are defined, and their potent binding activity with human DC-SIGN, a receptor associated with the gut lymphoid tissue, is disclosed.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-17992-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7434892PMC
August 2020

Unveiling Molecular Recognition of Sialoglycans by Human Siglec-10.

iScience 2020 Jun 2;23(6):101231. Epub 2020 Jun 2.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Via Cintia 4, 80126 Napoli, Italy. Electronic address:

Siglec-10 is an inhibitory I-type lectin selectively recognizing sialoglycans exposed on cell surfaces, involved in several patho-physiological processes. The key role Siglec-10 plays in the regulation of immune cell functions has made it a potential target for the development of immunotherapeutics against a broad range of diseases. However, the crystal structure of the protein has not been resolved for the time being and the atomic description of Siglec-10 interactions with complex glycans has not been previously unraveled. We present here the first insights of the molecular mechanisms regulating the interaction between Siglec-10 and naturally occurring sialoglycans. We used combined spectroscopic, computational and biophysical approaches to dissect glycans' epitope mapping and conformation upon binding in order to afford a description of the 3D complexes. Our outcomes provide a structural perspective for the rational design and development of high-affinity ligands to control the receptor functionality.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.isci.2020.101231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7306591PMC
June 2020

Structure of the unusual HH103 lipopolysaccharide and its role in symbiosis.

J Biol Chem 2020 08 16;295(32):10969-10987. Epub 2020 Jun 16.

Department of Microbiology, Faculty of Biology, University of Seville, Sevilla, Spain

Rhizobia are soil bacteria that form important symbiotic associations with legumes, and rhizobial surface polysaccharides, such as K-antigen polysaccharide (KPS) and lipopolysaccharide (LPS), might be important for symbiosis. Previously, we obtained a mutant of HH103, , that does not produce KPS, a homopolysaccharide of a pseudaminic acid derivative, but whose LPS electrophoretic profile was indistinguishable from that of the WT strain. We also previously demonstrated that the HH103 operon is responsible for 5-acetamido-3,5,7,9-tetradeoxy-7-(3-hydroxybutyramido)-l--l--nonulosonic acid [Pse5NAc7(3OHBu)] production and is involved in HH103 KPS and LPS biosynthesis and that an HH103 mutant cannot produce KPS and displays an altered LPS structure. Here, we analyzed the LPS structure of HH103 , focusing on the carbohydrate portion, and found that it contains a highly heterogeneous lipid A and a peculiar core oligosaccharide composed of an unusually high number of hexuronic acids containing β-configured Pse5NAc7(3OHBu). This pseudaminic acid derivative, in its α-configuration, was the only structural component of the HH103 KPS and, to the best of our knowledge, has never been reported from any other rhizobial LPS. We also show that Pse5NAc7(3OHBu) is the complete or partial epitope for a mAb, NB6-228.22, that can recognize the HH103 LPS, but not those of most of the strains tested here. We also show that the LPS from HH103 is identical to that of HH103 but devoid of any Pse5NAc7(3OHBu) residues. Notably, this mutant was severely impaired in symbiosis with its host, .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1074/jbc.RA120.013393DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7415993PMC
August 2020

Adaptive defence-related changes in the metabolome of Sorghum bicolor cells in response to lipopolysaccharides of the pathogen Burkholderia andropogonis.

Sci Rep 2020 05 6;10(1):7626. Epub 2020 May 6.

Research Centre for Plant Metabolomics, Department of Biochemistry, University of Johannesburg, Auckland Park, 2006, South Africa.

Plant cell suspension culture systems are valuable for the study of complex biological systems such as inducible defence responses and aspects of plant innate immunity. Perturbations to the cellular metabolome can be investigated using metabolomic approaches in order to reveal the underlying metabolic mechanism of cellular responses. Lipopolysaccharides from the sorghum pathogen, Burkholderia andropogonis (LPS), were purified, chemically characterised and structurally elucidated. The lipid A moiety consists of tetra- and penta-acylated 1,4'-bis-phosphorylated disaccharide backbone decorated by aminoarabinose residues, while the O-polysaccharide chain consists of linear trisaccharide repeating units of [→2)-α-Rha3CMe-(1 → 3)-α-Rha-(1 → 3)-α-Rha-(1 → ]. The effect of LPS in triggering metabolic reprogramming in Sorghum bicolor cells were investigated using untargeted metabolomics with liquid chromatography coupled to mass spectrometry detection. Cells were treated with LPS and the metabolic changes monitored over a 30 h time period. Alterations in the levels of phytohormones (jasmonates, zeatins, traumatic-, azelaic- and abscisic acid), which marked the onset of defence responses and accumulation of defence-related metabolites, were observed. Phenylpropanoids and indole alkaloids as well as oxylipins that included di- and trihydroxyoctadecedienoic acids were identified as signatory biomarkers, with marked secretion into the extracellular milieu. The study demonstrated that sorghum cells recognise LPS as a 'microbe-associated molecular pattern', perturbing normal cellular homeostasis. The molecular features of the altered metabolome were associated with phytohormone-responsive metabolomic reconfiguration of primary and secondary metabolites originating from various metabolic pathways, in support of defence and immunity.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-64186-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7203242PMC
May 2020

Structural basis for Glycan-receptor binding by mumps virus hemagglutinin-neuraminidase.

Sci Rep 2020 01 31;10(1):1589. Epub 2020 Jan 31.

Department of Chemical Sciences, Complesso Universitario Monte Sant'Angelo, University of Naples Federico II, Via Cintia 4, I-80126, Napoli, Italy.

Mumps virus is one of the main cause of respiratory illnesses in humans, especially children. Among the viral surface glycoproteins, the hemagglutinin - neuraminidase, MuV-HN, plays key roles in virus entry into host cells and infectivity, thus representing an ideal target for the design of novel inhibitors. Here we report the detailed analysis of the molecular recognition of host cell surface sialylated glycans by the viral glycoprotein MuV-HN. By a combined use of NMR, docking, molecular modelling and CORCEMA-ST, the structural features of sialoglycans/MuV-HN complexes were revealed. Evidence for a different enzyme activity toward longer and complex substrates compared to unbranched ligands was also examined by an accurate NMR kinetic analysis. Our results provide the basis for the structure-based design of effective drugs against mumps-induced diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-58559-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6994497PMC
January 2020

The Core Fucose on an IgG Antibody is an Endogenous Ligand of Dectin-1.

Angew Chem Int Ed Engl 2019 12 7;58(51):18697-18702. Epub 2019 Nov 7.

Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka, 560-0043, Japan.

The core fucose, a major modification of N-glycans, is implicated in immune regulation, such as the attenuation of the antibody-dependent cell-mediated cytotoxicity of antibody drugs and the inhibition of anti-tumor responses via the promotion of PD-1 expression on T cells. Although the core fucose regulates many biological processes, no core fucose recognition molecule has been identified in mammals. Herein, we report that Dectin-1, a known anti-β-glucan lectin, recognizes the core fucose on IgG antibodies. A combination of biophysical experiments further suggested that Dectin-1 recognizes aromatic amino acids adjacent to the N-terminal asparagine at the glycosylation site as well as the core fucose. Thus, Dectin-1 appears to be the first lectin-like molecule involved in the heterovalent and specific recognition of characteristic N-glycans on antibodies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/anie.201911875DOI Listing
December 2019

Synthesis of Forsythenethoside A, a Neuroprotective Macrocyclic Phenylethanoid Glycoside, and NMR Analysis of Conformers.

J Org Chem 2019 11 24;84(21):13733-13743. Epub 2019 Oct 24.

State Key Laboratory of Bioorganic and Natural Products Chemistry, Center for Excellence in Molecular Synthesis, Shanghai Institute of Organic Chemistry , University of Chinese Academy of Sciences, Chinese Academy of Sciences , 345 Lingling Road , Shanghai 200032 , China.

Forsythenethoside A () is a structurally unique macrocyclic phenylethanoid glycoside, which was isolated from and displayed considerable neuroprotective activities. Here, we report its first chemical synthesis via a longest linear sequence of 14 steps in 5% overall yield wherein intramolecular oxidative coupling was successfully employed to realize the pivotal macrocyclization. NMR analysis revealed the existence of an unexpected conformational interconversion of the congested macrocycles.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acs.joc.9b01956DOI Listing
November 2019

Weak Agonistic LPS Restores Intestinal Immune Homeostasis.

Mol Ther 2019 11 19;27(11):1974-1991. Epub 2019 Jul 19.

Institute of Medical Microbiology and Hygiene, University of Tübingen, Tübingen, Germany. Electronic address:

Generated by gram-negative bacteria, lipopolysaccharides (LPSs) are one of the most abundant and potent immunomodulatory substances present in the intestinal lumen. Interaction of agonistic LPS with the host myeloid-differentiation-2/Toll-like receptor 4 (MD-2/TLR4) receptor complex results in nuclear factor κB (NF-κB) activation, followed by the robust induction of pro-inflammatory immune responses. Here we have isolated LPS from a common gut commensal, Bacteroides vulgatus mpk (BVMPK), which provides only weak agonistic activity. This weak agonistic activity leads to the amelioration of inflammatory immune responses in a mouse model for experimental colitis, and it was in sharp contrast to strong agonists and antagonists. In this context, the administration of BVMPK LPS into mice with severe intestinal inflammation re-established intestinal immune homeostasis within only 2 weeks, resulting in the clearance of all symptoms of inflammation. These inflammation-reducing properties of weak agonistic LPS are grounded in the induction of a special type of endotoxin tolerance via the MD-2/TLR4 receptor complex axis in intestinal lamina propria CD11c cells. Thus, weak agonistic LPS represents a promising agent to treat diseases involving pathological overactivation of the intestinal immune system, e.g., in inflammatory bowel diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ymthe.2019.07.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6838991PMC
November 2019

Characterisation of the Dynamic Interactions between Complex N-Glycans and Human CD22.

Chembiochem 2020 01 17;21(1-2):129-140. Epub 2019 Oct 17.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Via Cintia 4, 80126, Napoli, Italy.

CD22 (Siglec-2) is a B-cell surface inhibitory protein capable of selectively recognising sialylated glycans, thus dampening autoimmune responses against self-antigens. Here we have characterised the dynamic recognition of complex-type N-glycans by human CD22 by means of orthogonal approaches including NMR spectroscopy, computational methods and biophysical assays. We provide new molecular insights into the binding mode of sialoglycans in complex with h-CD22, highlighting the role of the sialic acid galactose moieties in the recognition process, elucidating the conformational behaviour of complex-type N-glycans bound to Siglec-2 and dissecting the formation of CD22 homo-oligomers on the B-cell surface. Our results could enable the development of additional therapeutics capable of modulating the activity of h-CD22 in autoimmune diseases and malignancies derived from B-cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/cbic.201900295DOI Listing
January 2020

Human Macrophage Galactose-Type Lectin (MGL) Recognizes the Outer Core of Escherichia coli Lipooligosaccharide.

Chembiochem 2019 07 24;20(14):1778-1782. Epub 2019 Jun 24.

Department of Chemical Sciences, University of Naples Federico II, via Cintia 4, 80126, Napoli, Italy.

Carbohydrate-lectin interactions intervene in and mediate most biological processes, including a crucial modulation of immune responses to pathogens. Despite growing interest in investigating the association between host receptor lectins and exogenous glycan ligands, the molecular mechanisms underlying bacterial recognition by human lectins are still not fully understood. Herein, a novel molecular interaction between the human macrophage galactose-type lectin (MGL) and the lipooligosaccharide (LOS) of Escherichia coli strain R1 is described. Saturation transfer difference NMR spectroscopy analysis, supported by computational studies, demonstrated that MGL bound to the purified deacylated LOS mainly through recognition of its outer core and established crucial interactions with the terminal Galα(1,2)Gal epitope. These results assess the ability of MGL to recognise glycan moieties exposed on Gram-negative bacterial surfaces.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/cbic.201900087DOI Listing
July 2019

Lipopolysaccharide structures of Gram-negative populations in the gut microbiota and effects on host interactions.

FEMS Microbiol Rev 2019 05;43(3):257-272

Department of Chemical Sciences, University of Naples Federico II, via Cinthia 4, 80126 Naples, Italy.

The human gastrointestinal tract harbors a heterogeneous and complex microbial community, which plays a key role in human health. The gut microbiota controls the development of the immune system by setting systemic threshold for immune activation. Glycoconjugates, such as lipopolysaccharides, from gut bacteria have been shown to be able to elicit both systemic proinflammatory and immunomodulatory responses. This phenomenon is particularly intriguing considering that the immune system is charged with the task to distinguish the beneficial microbes from the pathogens, even if the commensal bacteria have molecular patterns resembling those of the pathogenic counterparts. Therefore, the importance of the chemical structure of these macromolecules in fine tuning this delicate equilibrium is beyond question. This review offers an overview of the current understanding of chemical peculiarities of the lipopolysaccharides isolated from the gut microbiota, and their relationships to their biological activity in terms of immune system maturation and development.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/femsre/fuz002DOI Listing
May 2019

Cancer Immunotherapy of TLR4 Agonist-Antigen Constructs Enhanced with Pathogen-Mimicking Magnetite Nanoparticles and Checkpoint Blockade of PD-L1.

Small 2019 01 20;15(4):e1803993. Epub 2018 Dec 20.

Department of Chemistry and Centre for NanoHealth, Swansea University, Singleton Park, Swansea, SA2 8PP, UK.

Despite the tremendous potential of Toll-like receptor 4 (TLR4) agonists in vaccines, their efficacy as monotherapy to treat cancer has been limited. Only some lipopolysaccharides (LPS) isolated from particular bacterial strains or structures like monophosphoryl lipid A (MPLA) derived from lipooligosaccharide (LOS), avoid toxic overactivation of innate immune responses while retaining adequate immunogenicity to act as adjuvants. Here, different LOS structures are incorporated into nanoparticle-filled phospholipid micelles for efficient vaccine delivery and more potent cancer immunotherapy. The structurally unique LOS of the plant pathogen Xcc is incorporated into phospholipid micelles encapsulating iron oxide nanoparticles, producing stable pathogen-mimicking nanostructures suitable for targeting antigen presenting cells in the lymph nodes. The antigen is conjugated via a hydrazone bond, enabling rapid, easy-to-monitor and high-yield antigen ligation at low concentrations. The protective effect of these constructs is investigated against a highly aggressive model for tumor immunotherapy. The results show that the nanovaccines lead to a higher-level antigen-specific cytotoxic T lymphocyte (CTL) effector and memory responses, which when combined with abrogation of the immunosuppressive programmed death-ligand 1 (PD-L1), provide 100% long-term protection against repeated tumor challenge. This nanovaccine platform in combination with checkpoint inhibition of PD-L1 represents a promising approach to improve the cancer immunotherapy of TLR4 agonists.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/smll.201803993DOI Listing
January 2019

Zymomonas mobilis exopolysaccharide structure and role in high ethanol tolerance.

Carbohydr Polym 2018 Dec 20;201:293-299. Epub 2018 Aug 20.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Via Cintia 4, I-80126 Napoli, Italy. Electronic address:

Bacterial cell surface exopolysaccharides (EPS) provide a protective barrier from the external milieu and participate in host-environment interactions. Zymomonas mobilis, an ethanologenic Gram negative bacterium, is used by the industry in bio-ethanol production process, due to its extraordinary resistance to a highly ethanolic environment. We found that Z. mobilis produces a mixture of two EPSs, an [α-(1→6)-D-Manp] mannose homopolymer and a galactose containing polysaccharide: [→2)-β-D-Galf-(1→3)-β-D-Galp-(1→]. A physico-chemical study, conducted with diffusion-ordered spectroscopy (DOSY) and Dynamic Light Scattering (DLS), allowed to demonstrate that, differently from the majority of polysaccharides, ethanol is a good solvent for the galactose containing EPS, revealing that its chemical structure is specifically connected with the Zymomonas mobilis high ethanol tolerance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.carbpol.2018.08.072DOI Listing
December 2018

The Lipid A Structure from the Marine Sponge Symbiont Endozoicomonas sp. HEX 311.

Chembiochem 2019 01 20;20(2):230-236. Epub 2018 Nov 20.

Department of Chemical Sciences, University of Naples Federico II, via Cinthia 4, 80126, Naples, Italy.

Endozoicomonas sp. HEX311 is a Gram-negative bacterium known to establish a commensal interaction with the marine demosponge Suberites domuncula. The molecular bases of the sponge-microbe interaction events are still poorly defined. Nevertheless, it has been proved that S. domuncula possesses an innate immune system with similarities to the mammalian one and is able to recognize the main component of the Gram-negative bacteria cell wall: the lipopolysaccharide. Whether this recognition occurs in a structure-dependent manner, which is typical for mammalian immune system receptors, is still under investigation. Herein, we report the Endozoicomonas sp. HEX311 lipid A structure obtained by a combination of data attained from chemical, MALDI MS, and MS approaches. The lipid A is a complex family of species decorated by pyrophosphate and phosphate units and carrying (R)-3-hydroxydodecanoic acid, (R)-3-hydroxytetradecanonic acid, iso-2-hydroxyundecanoic acid, iso-(R)-3-hydroxyundecanoic acid, and iso-nonanoic acid as acyl chains.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/cbic.201800441DOI Listing
January 2019

Lipid A: Immunological Properties and Molecular Basis of Its Binding to the Myeloid Differentiation Protein-2/Toll-Like Receptor 4 Complex.

Front Immunol 2018 14;9:1888. Epub 2018 Aug 14.

Dipartimento di Scienze Chimiche, Complesso Universitario Monte Sant'Angelo, Università di Napoli Federico II, Naples, Italy.

Lipopolysaccharides (LPS) are potent activator of the innate immune response through the binding to the myeloid differentiation protein-2 (MD-2)/toll-like receptor 4 (TLR4) receptor complexes. Although a variety of LPSs have been characterized so far, a detailed molecular description of the structure-activity relationship of the lipid A part has yet to be clarified. Photosynthetic strains, symbiont of legumes, express distinctive LPSs bearing very long-chain fatty acids with a hopanoid moiety covalently linked to the lipid A region. Here, we investigated the immunological properties of LPSs isolated from strains on both murine and human immune systems. We found that they exhibit a weak agonistic activity and, more interestingly, a potent inhibitory effect on MD-2/TLR4 activation exerted by toxic enterobacterial LPSs. By applying computational modeling techniques, we also furnished a plausible explanation for the LPS inhibitory activity at atomic level, revealing that its uncommon lipid A chemical features could impair the proper formation of the receptorial complex, and/or has a destabilizing effect on the pre-assembled complex itself.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2018.01888DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6102379PMC
September 2019