Angelina Sá Palma

Universidade NOVA de Lisboa | NOVA · Department of Chemistry

Glycan microarrays are essential tools in glycobiology and are being widely used for assignment of glycan ligands in diverse glycan recognition systems. We have developed a new software, called Carbohydrate microArray Analysis and Reporting Tool (CarbArrayART), to address the need for a distributable application for glycan microarray data managemen...

The primary recognition event between a fungal pathogen and the immune system normally involves the engagement of a pattern recognition receptor with specific components of the cell wall. However, the cell wall is a complex three dimensional structure whose composition changes rapidly in accordance with environmental stimuli. Therefore it is import...

With the knowledge of bacterial gene systems encoding proteins that target dietary carbohydrates as a source of nutrients and their importance for human health, major efforts are being made to understand carbohydrate recognition by various commensal bacteria. Here, we describe an integrative strategy that combines carbohydrate microarray technology...

Glycan microarrays have played important roles in detection and specificity assignment of glycan-recognition by proteins. However, the size and diversity of glycan libraries in current microarray systems are small compared to estimated glycomes, and these may lead to missed detection or incomplete assignment. For microarray construction, covalent a...

Glycan microarrays have played important roles in detection and specificity assignment of glycan-recognition by proteins. However, the size and diversity of glycan libraries in current microarray systems are small compared to estimated glycomes, and these may lead to missed detection or incomplete assignment. For microarray construction, covalent a...

The immune microenvironment of breast cancer (BC) is composed by high macrophage infiltrates, correlated with the most aggressive subtypes. Tumour-associated macrophages (TAM) within the BC microenvironment are key regulators of immune suppression and BC progression. Nevertheless, several key questions regarding TAM polarisation by BC are still not...

The structural diversity of the lipopolysaccharides (LPSs) from Helicobacter pylori poses a challenge to establish accurate and strain-specific structure-function relationships in interactions with the host. Here, LPS structural domains from five clinical isolates were obtained and compared with the reference strain 26695. This was achieved combini...

Siglec-15 is a conserved sialic acid-binding Ig-like lectin expressed on osteoclast progenitors that plays an important role in osteoclast development and function. It is also expressed by tumor-associated macrophages and by some tumors, where it is thought to contribute to the immunosuppressive microenvironment. It was shown previously that engage...

During the course of fungal infection, pathogen recognition by the innate immune system is critical to initiate efficient protective immune responses. The primary event that triggers immune responses is the binding of Pattern Recognition Receptors (PRRs), which are expressed at the surface of host immune cells, to Pathogen-Associated Molecular Patt...

Understanding the specific molecular interactions between proteins and β1,3‐1,4‐mixed‐linked D‐glucans is fundamental to harvest the full biological and biotechnological potential of these carbohydrates and of proteins that specifically recognise them. The family 11 carbohydrate‐binding module from Clostridium thermocellum (CtCBM11) is known for it...

The human natural killer-1 (HNK-1) epitope is a unique sulfated trisaccharide sequence presented on O- and N-glycans of various glycoproteins and on glycolipids. It is overexpressed in the nervous system and plays crucial roles in nerve regeneration, synaptic plasticity, and neuronal diseases. However, the investigation of functional roles of HNK-1...

The emergence of bacterial resistance to different antibiotics in clinical use, together with the knowledge on the mechanisms by which bacteria resist the action of aminoglycosides, have contributed to the renewed interest in these molecules as potential antimicrobials. Here, we give an overview on natural and semisynthetic aminoglycosides and thei...

During the course of fungal infection, pathogen recognition by the innate immune system is critical to initiate efficient protective immune responses. The primary event that triggers immune responses is the binding of Pattern Recognition Receptors (PRRs), which are expressed at the surface of host immune cells, to Pathogen-Associated Molecular Patt...

The relevance of microalgae biotechnology for producing high-value compounds with biomedical application, such as polysaccharides, has been increasing. Despite this, the knowledge about the composition and structure of microalgae polysaccharides is still scarce. In this work, water-soluble polysaccharides from Nannochloropsis oculata were extracted...

The study of glycan recognition by glycan-binding proteins has emerged as a challenging area of research at the interface of chemistry and biology. The advances in glycomic approaches, including the development and successful application of glycan microarrays, have played a crucial role in the understanding of glycan functions. A variety of methods...

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The high global burden of over one million annual lethal fungal infections reflects a lack of protective vaccines, late diagnosis and inadequate chemotherapy. Here, we have generated a unique set of fully human anti-Candida monoclonal antibodies (mAbs) with diagnostic and therapeutic potential by expressing recombinant antibodies from genes cloned...

Incomplete O-glycosylation is a feature associated with malignancy resulting in the expression of truncated glycans such as the sialyl-Tn (STn) antigen. Despite all the progress in the development of potential anti-cancer antibodies, their application is frequently hindered by low specificities and cross-reactivity. In this study, a novel anti-STn...

O-glycosylation is a post-translational modification of proteins crucial to molecular mechanisms in health and disease. O-glycans are typically highly heterogeneous. The involvement of specific O-glycan sequences in many bio-recognition systems is yet to be determined due to a lack of efficient methodologies. We describe here a targeted microarray...

Glucans are polysaccharides of increasing biomedical interest because of their involvement in mechanisms of pathogen recognition, modulation of the immune system and anticancer, and health-promoting activities. Most of these biological activities occur through specific interactions with glucan-recognizing proteins. However, detailed molecular studi...

MIRAGE (Minimum Information Required for A Glycomics Experiment) is an initiative that was created by experts in the fields of glycobiology, glycoanalytics and glycoinformatics to produce guidelines for reporting results from the diverse types of experiments and analyses used in structural and functional studies of glycans in the scientific literat...

The β1,2-glucans produced by bacteria are important in invasion, survival and immunomodulation in infected hosts be they mammals or plants. However, there has been a lack of information on proteins which recognize these molecules. This is partly due to the extremely limited availability of the sequence-defined oligosaccharides and derived probes fo...

Glucans are polymers of D-glucose with differing linkages in linear or branched sequences. They are constituents of microbial and plant cell-walls and involved in important bio-recognition processes including immunomodulation, anti-cancer activities, pathogen virulence and plant cell-wall biodegradation. Translational possibilities for these activi...

For a subset of pathogenic microorganisms, including Streptococcus pneumoniae, the recognition and degradation of host hyaluronan contributes to bacterial spreading through the extracellular matrix and enhancing access to host cell surfaces. The hyaluronate lyase (Hyl) presented on the surface of S. pneumoniae performs this role. Using glycan micro...

Monoclonal antibody F77 was previously raised against human prostate cancer cells and has been shown to recognize a carbohydrate antigen, but the carbohydrate sequence of the antigen was elusive. Here, we make multifaceted approaches to characterize F77 antigen, including binding analyses with the glycolipid extract of the prostate cancer cell line...

The neoglycolipid (NGL) technology is the basis of a state-of-the-art oligosaccharide microarray system. The NGL-based microarray system in the Glycosciences Laboratory Imperial College London (http://www3.imperial.ac.uk/glycosciences) is one of the two leading platforms for glycan microarrays, being offered for screening analyses to the broad biom...

Viruses within a family often vary in their cellular tropism and pathogenicity. In many cases, these variations are due to viruses switching their specificity from one cell surface receptor to another. The structural requirements that underlie such receptor switching are not well understood especially for carbohydrate-binding viruses, as methods ca...

B-Lymphotropic Polyomavirus (LPyV) serves as a paradigm of virus receptor binding and tropism, and is the closest relative of the recently discovered Human Polyomavirus 9 (HPyV9). LPyV infection depends on sialic acid on host cells, but the molecular interactions underlying LPyV-receptor binding were unknown. We find by glycan array screening that...

As avian influenza A(H5N1) viruses continue to circulate in Asia and Africa, global concerns of an imminent pandemic persist. Recent experimental studies suggest that efficient transmission between humans of current H5N1 viruses only requires a few genetic changes. An essential step is alteration of the virus hemagglutinin from preferential binding...

In this chapter, we describe the key steps of the "designer" oligosaccharide microarray approach we followed to prove the carbohydrate binding activity and define the oligosaccharide ligands for Dectin-1, an atypical C-type lectin-like signaling receptor of the mammalian innate immune system with a key role in anti-fungal immunity. The term "design...

Carbohydrate microarrays, since their advent in 2002, are revolutionizing studies of the molecular basis of protein-carbohydrate interactions both in endogenous recognition systems and pathogen-host interactions. We have developed a unique carbohydrate microarray system based on the neoglycolipid (NGL) technology, a well-validated microscale approa...

Binding of individual MARR of EtMIC3 to fixed MDBK cell monolayers determined by ELISA. (PDF)

Chemical shift mapping for the interaction of recombinant EtMIC3-MAR1b with α2–6 sialyllactose. (Top) 1H-15N HSQC spectrum for 15N,13C-labelled EtMIC3-MARb alone (black) and in presence of unlabelled Siaα2–6Galβ1–4Glc (pink) at a molar ratio of 1∶1. (Bottom) Surface representation for the lowest energy structure for EtMIC3-MARb with residues colour...

Oligosaccharide probes included in the microarrays sorted by sialyl linkage and backbone sequences, and the binding signals (fluorescence intensities) they elicited* with EtMIC3-MAR5 and TgMIC1-MARR. (PDF)

Inhibition of Cell binding of EtMIC3-MAR5. Recombinant expressed protein (10 µg/ml) was incubated with either (1) fetuin -100 µg/ml (2) sialic acid -100 µgml (3) trisialoganglioside GT1a -100 µg/ml (4) disialoganglioside GD1a (Sigma) -100 µg/ml (5) disialoganglioside GD1b(Sigma) - 100 µg/ml, (6) disialoganglioside GD1a (Alexis biochemicals) -100 µg...

Three-dimensional structure of the MAR1b domain from EtMIC3. a) Stereo-view of the superimposition for the ten best NMR structures of EtMIC3-MAR1b. b) Superposition of EtMIC3-MAR1b (red; PDB code 2LBO) on the MAR2 domain from TgMIC1 (PDB code 2JH1; cyan). c) Superposition of EtMIC3-MAR1b (red; PDB code 2LBO) on the MAR1 domain from TgMIC1 (PDB code...

EtMIC3 Topology and MAR2 domain sequence alignment (Top) Cartoon shows the location of each type of MARR in the full length EtMIC3 protein. (Bottom) Sequence alignments for the MAR domain families from EtMIC3 and TgMIC1. Cysteines are shaded orange and disulfide bond connectivities are indicated for the MAR domains. The position of the MAR1 inserti...

Intermolecular NOE restraints for HADDOCK docking calculations of EtMIC3 with sialyl carbohydrates. (PDF)

Cumulative faecal oocysts counts from each individual bird in the two vaccination/challenge experiments shown in Figure 7 . (PDF)

Eimeria spp. are a highly successful group of intracellular protozoan parasites that develop within intestinal epithelial cells of poultry, causing coccidiosis. As a result of resistance against anticoccidial drugs and the expense of manufacturing live vaccines, it is necessary to understand the relationship between Eimeria and its host more deeply...

There is an increasing interest in the development of therapeutic antibodies (Ab) to improve the control of fungal pathogens, but none of these reagents is available for clinical use. We previously described a murine monoclonal antibody (mAb 2G8) targeting β-glucan, a cell wall polysaccharide common to most pathogenic fungi, which conferred signifi...

High-resolution X-ray crystallography and NMR studies by Koharudin and Gronenborn in this issue provide new information on the mode of N-glycan recognition by a cyanobacterial agglutinin, with anti-HIV activity pointing to the pentamannosyl core as a novel target for therapeutic intervention.

The human C-type lectin-like molecule CLEC5A is a critical macrophage receptor for dengue virus. The binding of dengue virus to CLEC5A triggers signaling through the associated adapter molecule DAP12, stimulating proinflammatory cytokine release. We have crystallized an informative ensemble of CLEC5A structural conformers at 1.9-Å resolution and de...

The human C-type lectin-like molecule CLEC5A is a critical macrophage receptor for dengue virus. The binding of dengue virus to CLEC5A triggers signaling through the associated adapter molecule DAP12, stimulating proinflammatory cytokine release. We have crystallized an informative ensemble of CLEC5A structural conformers at 1.9-Å resolution and de...

The term human epithelial carcinoma antigen (HCA) has been applied collectively to mucin-type high molecular weight (>1000kDa) glycoproteins that are over-expressed in epithelial cancers. Since the 1990s, over 40 monoclonal antibodies have been raised that recognize HCA. There has been evidence that the antigenic determinants are mostly carbohydrat...

The human C-type lectin-like molecule CLEC5A is a critical macrophage receptor for dengue virus. The binding of dengue virus to CLEC5A triggers signaling through the associated adapter molecule DAP12, stimulating proinflammatory cytokine release. We have crystallized an informative ensemble of CLEC5A structural conformers at 1.9-Å resolution and de...

Calreticulin is a chaperone of the endoplasmic reticulum (ER) assisting proteins in achieving the correctly folded structure. Details of the binding specificity of calreticulin are still a matter of debate. Calreticulin has been described as an oligosaccharide-binding chaperone but data are also accumulating in support of calreticulin as a polypept...

In this chapter, we describe the key procedures for isolation of the oligosaccharides and the preparation of neoglycolipid probes together with expression of malectin that have enabled the discovery of the highly selective binding of this newly described protein in the endoplasmic reticulum (ER) to a diglucosyl high-mannose N-glycan. This is the fi...

Mutations in the receptor-binding site of the hemagglutinin of pandemic influenza A(H1N1) 2009 viruses have been detected sporadically. An Asp222Gly (D222G) substitution has been associated with severe or fatal disease. Here we show that 222G variants infected a higher proportion of ciliated cells in cultures of human airway epithelium than did vir...

The human JC polyomavirus (JCV) causes a fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML), in immunocompromised individuals. Current treatment options for PML are inadequate. Sialylated oligosaccharides and the serotonin receptor are known to be necessary for JCV entry, but the molecular interactions underlying JCV atta...

Numerous intracellular pathogens exploit cell surface glycoconjugates for host cell recognition and entry. Unlike bacteria and viruses, Toxoplasma gondii and other parasites of the phylum Apicomplexa actively invade host cells, and this process critically depends on adhesins (microneme proteins) released onto the parasite surface from intracellular...

Carbohydrate chains of glycoproteins, glycolipids, proteoglycans, and polysaccharides mediate processes of biological and medical importance through their interactions with complementary proteins. The unraveling of these interactions is therefore a priority in biomedical sciences. Carbohydrate microarray technology is a new development at the front...

SDS-PAGE and Western blot analysis of IgG mAb-reactive material purified by immunoaffinity from hyphal secretion. Lane 1 and 2 show silver staining of the total hyphal secretion and the IgG mAb-immunopurified fraction, respectively. Lane 3 and 4 shows Western blot reactivity of the immunopurified fraction with the IgG mAb (Lane 3) or with the IgG f...

Concanavalin A- and IgG mAb-staining of hyphal secretion before and after periodate oxidation. Fungal secretions were treated 30 min with 0.1 M sodium periodate, dyalised by gel filtration and then compared to untreated secretion by SDS-PAGE and Western blot, followed by specific mannoprotein staining with digoxigenin-labelled Concanavalin A (Con-A...