Biochemical characterization of a SusD-like protein involved in β-1,3-glucan utilization by an uncultured cow rumen .

In ruminants, the rumen is a specialized stomach that is adapted to the breakdown of plant-derived complex polysaccharides through the coordinated activities of a diverse microbial community. Bacteroidota is a major phylum in this bovine rumen microbiota. They contain several clusters of genes called polysaccharide utilization loci (PULs) that encode proteins working in concert to capture, degrade, and transport polysaccharides.

Structural and Biochemical Characterization of a Nonbinding SusD-Like Protein Involved in Xylooligosaccharide Utilization by an Uncultured Human Gut Strain.

In the human gut microbiota, break down dietary and endogenous glycosides through highly specific polysaccharide utilization loci (PULs). PULs encode a variety of sensor regulators, binding proteins, transporters, and carbohydrate-active enzymes (CAZymes). Surface glycan-binding proteins (SGBPs) are essential for the efficient capture of the glycosides present on the cell surface, providing with a competitive advantage in colonizing their habitats.

Discovery and Biotechnological Exploitation of Glycoside-Phosphorylases.

Among carbohydrate active enzymes, glycoside phosphorylases (GPs) are valuable catalysts for white biotechnologies, due to their exquisite capacity to efficiently re-modulate oligo- and poly-saccharides, without the need for costly activated sugars as substrates. The reversibility of the phosphorolysis reaction, indeed, makes them attractive tools for glycodiversification. However, discovery of new GP functions is hindered by the difficulty in identifying them in sequence databases, and, rather, relies on extensive and tedious biochemical characterization studies.

Bacterial α-Glucan and Branching Sucrases from GH70 Family: Discovery, Structure-Function Relationship Studies and Engineering.

Glucansucrases and branching sucrases are classified in the family 70 of glycoside hydrolases. They are produced by lactic acid bacteria occupying very diverse ecological niches (soil, buccal cavity, sourdough, intestine, dairy products, etc.).

hnRNP H/F drive RNA G-quadruplex-mediated translation linked to genomic instability and therapy resistance in glioblastoma.

RNA G-quadruplexes (RG4s) are four-stranded structures known to control mRNA translation of cancer relevant genes. RG4 formation is pervasive in vitro but not in cellulo, indicating the existence of poorly characterized molecular machinery that remodels RG4s and maintains them unfolded. Here, we performed a quantitative proteomic screen to identify cytosolic proteins that interact with a canonical RG4 in its folded and unfolded conformation.

A specific oligosaccharide-binding site in the alternansucrase catalytic domain mediates alternan elongation.

Microbial α-glucans produced by GH70 (glycoside hydrolase family 70) glucansucrases are gaining importance because of the mild conditions for their synthesis from sucrose, their biodegradability, and their current and anticipated applications that largely depend on their molar mass. Focusing on the alternansucrase (ASR) from NRRL B-1355, a well-known glucansucrase catalyzing the synthesis of both high- and low-molar-mass alternans, we searched for structural traits in ASR that could be involved in the control of alternan elongation. The resolution of five crystal structures of a truncated ASR version (ASRΔ2) in complex with different gluco-oligosaccharides pinpointed key residues in binding sites located in the A and V domains of ASR.

Processivity of dextransucrases synthesizing very-high-molar-mass dextran is mediated by sugar-binding pockets in domain V.

The dextransucrase DSR-OK from the Gram-positive bacterium DSM17330 produces a dextran of the highest molar mass reported to date (∼10 g/mol). In this study, we selected a recombinant form, DSR-OKΔ1, to identify molecular determinants involved in the sugar polymerization mechanism and that confer its ability to produce a very-high-molar-mass polymer. In domain V of DSR-OK, we identified seven putative sugar-binding pockets characteristic of glycoside hydrolase 70 (GH70) glucansucrases that are known to be involved in glucan binding.

Futile Encounter Engineering of the DSR-M Dextransucrase Modifies the Resulting Polymer Length.

The factors that define the resulting polymer length of distributive polymerases are poorly understood. Here, starting from the crystal structure of the dextransucrase DSR-M in complex with an isomaltotetraose, we define different anchoring points for the incoming acceptor. Mutation of one of these, Trp624, decreases the catalytic rate of the enzyme but equally skews the size distribution of the resulting dextran chains toward shorter chains.

C1q and Mannose-Binding Lectin Interact with CR1 in the Same Region on CCP24-25 Modules.

Complement receptor type 1 (CR1) is a multi modular membrane receptor composed of 30 homologous complement control protein modules (CCP) organized in four different functional regions called long homologous repeats (LHR A, B, C, and D). CR1 is a receptor for complement-opsonins C3b and C4b and specifically interacts through pairs of CCP modules located in LHR A, B, and C. Defense collagens such as mannose-binding lectin (MBL), ficolin-2, and C1q also act as opsonins and are involved in immune clearance through binding to the LHR-D region of CR1.

Conferring cellulose-degrading ability to to facilitate a consolidated bioprocessing approach.

Background: , one of the most widely studied "nonconventional" oleaginous yeast species, is unable to grow on cellulose. Recently, we identified and overexpressed two endogenous β-glucosidases in , thus enabling this yeast to use cello-oligosaccharides as a carbon source for growth. Using this engineered yeast platform, we have now gone further toward building a fully cellulolytic for use in consolidated bioprocessing of cellulose.

Structures of parasite calreticulins provide insights into their flexibility and dual carbohydrate/peptide-binding properties.

Calreticulin (CRT) is a multifaceted protein, initially discovered as an endoplasmic reticulum (ER) chaperone protein, that is essential in calcium metabolism. Various implications in cancer, early development and immunology have been discovered more recently for CRT, as well as its role as a dominant 'eat-me' prophagocytic signal. Intriguingly, cell-surface exposure/secretion of CRT is among the infective strategies used by parasites such as , , , and .

Probing the Functions of Carbohydrate Binding Modules in the CBEL Protein from the Oomycete Phytophthora parasitica.

Oomycetes are microorganisms that are distantly related to true fungi and many members of this phylum are major plant pathogens. Oomycetes express proteins that are able to interact with plant cell wall polysaccharides, such as cellulose. This interaction is thought to be mediated by carbohydrate-binding modules that are classified into CBM family 1 in the CAZy database.

Development of cellobiose-degrading ability in Yarrowia lipolytica strain by overexpression of endogenous genes.

Background: Yarrowia lipolytica, one of the most widely studied "nonconventional" oleaginous yeast species, is unable to grow on cellobiose. Engineering cellobiose-degrading ability into this yeast is a vital step towards the development of cellulolytic biocatalysts suitable for consolidated bioprocessing.

Results: In the present work, we identified six genes encoding putative β-glucosidases in the Y.

Structural bases for N-glycan processing by mannoside phosphorylase.

The first crystal structure of Uhgb_MP, a β-1,4-mannopyranosyl-chitobiose phosphorylase belonging to the GH130 family which is involved in N-glycan degradation by human gut bacteria, was solved at 1.85 Å resolution in the apo form and in complex with mannose and N-acetylglucosamine. SAXS and crystal structure analysis revealed a hexameric structure, a specific feature of GH130 enzymes among other glycoside phosphorylases.

Structural insights into Aspergillus fumigatus lectin specificity: AFL binding sites are functionally non-equivalent.

The Aspergillus fumigatus lectin AFL was recently described as a new member of the AAL lectin family. As a lectin from an opportunistic pathogen, it might play an important role in the interaction of the pathogen with the human host. A detailed study of structures of AFL complexed with several monosaccharides and oligosaccharides, including blood-group epitopes, was combined with affinity data from SPR and discussed in the context of previous findings.

A soluble fucose-specific lectin from Aspergillus fumigatus conidia--structure, specificity and possible role in fungal pathogenicity.

Aspergillus fumigatus is an important allergen and opportunistic pathogen. Similarly to many other pathogens, it is able to produce lectins that may be involved in the host-pathogen interaction. We focused on the lectin AFL, which was prepared in recombinant form and characterized.

Burkholderia cenocepacia lectin A binding to heptoses from the bacterial lipopolysaccharide.

Bacteria from the Burkholderia cepacia complex (Bcc) cause highly contagious pneumonia among cystic fibrosis (CF) patients. Among them, Burkholderia cenocepacia is one of the most dangerous in the Bcc and is the most frequent cause of morbidity and mortality in CF patients. Indeed, it is responsible of "cepacia syndrome", a deadly exacerbation of infection, that is the main cause of poor outcomes in lung transplantation.

Burkholderia cenocepacia BC2L-C is a super lectin with dual specificity and proinflammatory activity.

Lectins and adhesins are involved in bacterial adhesion to host tissues and mucus during early steps of infection. We report the characterization of BC2L-C, a soluble lectin from the opportunistic pathogen Burkholderia cenocepacia, which has two distinct domains with unique specificities and biological activities. The N-terminal domain is a novel TNF-α-like fucose-binding lectin, while the C-terminal part is similar to a superfamily of calcium-dependent bacterial lectins.

Low-temperature neutron diffraction structures of N-glycoprotein linkage models and analogues: structure refinement and trifurcated hydrogen bonds.

The biological addition of oligosaccharide moieties to asparagine residues of N-glycoproteins influences the properties and bioactivities of these macromolecules. The low-temperature neutron crystal structures of three N-glycoprotein linkage models and analogues provide accurate characterization of the three-dimensional structure of the conserved GlcNAc-Asn linkage. These first crystal structures of N-acetylated carbohydrates obtained by neutron diffraction provide high-resolution geometrical parameters that can be used for force-field parametrization and subsequent molecular dynamics simulation of N-glycoproteins.

A TNF-like trimeric lectin domain from Burkholderia cenocepacia with specificity for fucosylated human histo-blood group antigens.

The opportunistic pathogen Burkholderia cenocepacia expresses several soluble lectins, among them BC2L-C. This lectin exhibits two domains: a C-terminal domain with high sequence similarity to the recently described calcium-dependent mannose-binding lectin BC2L-A, and an N-terminal domain of 156 amino acids without similarity to any known protein. The recombinant N-terminal BC2L-C domain is a new lectin with specificity for fucosylated human histo-blood group epitopes H-type 1, Lewis b, and Lewis Y, as determined by glycan array and isothermal titration calorimetry.

Structures of the tumor necrosis factor alpha inducing protein Tipalpha: a novel virulence factor from Helicobacter pylori.

Helicobacter pylori secretes a unique virulence factor, Tipalpha, that enters gastric cells and both stimulates the production of the TNF-alpha and activates the NF-kappaB pathway. The structures of a truncated version of Tipalpha (TipalphaN34) in two crystal forms are presented here. Tipalpha adopts a novel beta(1)alpha(1)alpha(2)beta(2)beta(3)alpha(3)alpha(4) topology that can be described as a combination of three domains.

High affinity interaction between a bivalve C-type lectin and a biantennary complex-type N-glycan revealed by crystallography and microcalorimetry.

Codakine is an abundant 14-kDa mannose-binding C-type lectin isolated from the gills of the sea bivalve Codakia orbicularis. Binding studies using inhibition of hemagglutination indicated specificity for mannose and fucose monosaccharides. Further experiments using a glycan array demonstrated, however, a very fine specificity for N-linked biantennary complex-type glycans.

Unusual entropy-driven affinity of Chromobacterium violaceum lectin CV-IIL toward fucose and mannose.

The purple pigmented bacterium Chromobacterium violaceum is a dominant component of tropical soil microbiota that can cause rare but fatal septicaemia in humans. Its sequenced genome provides insight into the abundant potential of this organism for biotechnological and pharmaceutical applications and allowed an ORF encoding a protein that is 60% identical to the fucose binding lectin (PA-IIL) from Pseudomonas aeruginosa and the mannose binding lectin (RS-IIL) from Ralstonia solanacearum to be identified. The lectin, CV-IIL, has recently been purified from C.