Ana Lorena Morales-García

Soap Lake, a small meromictic lake in Washington State, USA, harbours a diverse microbial community adapted to high carbonate, sulfate and sulfide concentrations and a pH of approximately 9.8. Alkalitalea saponilacus , an anaerobic obligate bacterium from the lowest layer of Soap Lake, encodes multiple secreted enzymes, bioinformatically annotated as active on a range of carbohydrates. The ability of these enzymes to function under their extreme native conditions offers considerable industrial potential, most notably in the production of granular laundry and automatic dishwashing detergents that require robust performance under highly ionic conditions. Whilst some alkaline‐stable enzyme classes exist within current detergent formulations, there is a significant underrepresentation of non‐starch polysaccharide‐targeting candidates, positioning A. saponilacus as an untapped reservoir for this application. This study investigates A. saponilacus beta‐glucan degradation and confirms, using RT‐qPCR, the involvement of polysaccharide utilisation loci (PUL) 10 in this process. A representative glycoside hydrolase 16 from this PUL has been heterologously expressed and evaluated as a beta‐glucan‐targeting candidate, stable under harsh alkaline conditions. To demonstrate the industrial potential of A. saponilacus across both genomic location and functional enzyme families, a mannan‐targeting glycoside hydrolase 26 and a cellulose‐targeting glycoside hydrolase 9 were also evaluated for substrate deconstruction under high pH conditions. All three model GHs were tested for complex stain removal within a commercial detergent formula, where they exhibited considerable potential as additives. These results reinforce the prospects of the respective molecules, and ultimately the A. saponilacus proteome, for biotechnological applications where performance under alkaline or strongly ionic conditions is required.

β-mannans are plant structural and storage polysaccharides prevalent in the human diet. Their degradation in the gastrointestinal tract is mediated by the human gut microbiota (HGM) through expression of a plethora of carbohydrate-active enzymes (CAZymes), although our understanding of the details of mannan breakdown is lacking. In this study, a prominent HGM member, Bacteroides cellulosilyticus , was found to be exceptionally efficient at utilising β-mannans, mediated by the expression of a single polysaccharide utilisation locus (PUL). Amongst the predicted surface CAZymes encoded in the PUL, we identified a family 26 glycoside hydrolase of an unusual molecular architecture. Bc WH2_GH26 contains a putative carbohydrate-binding module (CBM) directly intercalated into its catalytic domain, unlike classical CBMs which are located at the N- or C-termini of the catalytic domain. Phylogenetic and functional analyses of this internal CBM, and a homologue from another mannan user Bacteroides uniformis , revealed a narrow specificity for β-mannan and support their classification as a novel CBM family. To investigate the evolutionary basis for the unusual enzyme architecture, the effect of the CBM on the catalytic activity of the enzyme was assessed. No significant differences in the kinetic parameters were found between the full-length and CBM deletion constructs against both soluble and insoluble mannans. The potential role of the internal CBM in enzyme function is discussed in the context of the likely localisation of the Bc WH2_GH26 in the outer membrane utilisome encoded by the Bc mannan PUL.

Attachment assays of a Pseudomonas isolate to fused silica slides showed that treatment with DNaseI significantly inhibited cellular adsorption, which was restored upon DNA treatment. These assays confirmed the important role of extracellular DNA (eDNA) adsorption to a surface. To investigate the eDNA adsorption mechanism, single-molecule force spectroscopy (SMFS) was used to measure the adsorption of eDNA to silicon surfaces in the presence of different concentrations of sodium and calcium ions. SMFS reveals that the work of adhesion required to remove calcium-bound eDNA from the silicon oxide surface is substantially greater than that for sodium. Molecular dynamics simulations were also performed, and here, it was shown that the energy gain in eDNA adsorption to a silicon oxide surface in the presence of calcium ions is small and much less than that in the presence of sodium. The simulations show that the length scales involved in eDNA adsorption are less in the presence of sodium ions than those in the presence of calcium. In the presence of calcium, eDNA is pushed above the surface cations, whereas in the presence of sodium ions, short-range interactions with the surface dominate. Moreover, SMFS data show that increasing [Ca²⁺] from 1 to 10 mM increases the adsorption of the cations to the silicon oxide surface and consequently enhances the Stern layer, which in turn increases the length scale associated with eDNA adsorption.

Direct glycan–glycan interactions are increasingly implicated in survival and pathogenicity of bacteria. Here, we show that they can be exploited by protozoan parasites in their insect hosts. Force spectroscopy revealed that Leishmania promastigotes display a high-affinity biomolecular interaction between their lipophosphoglycan glycocalyx and mimics of N-acetyl-D-galactosamine, commonly expressed on the midguts of a wide range of sand fly vector species. This enabled gut-adhesive nectomonad promastigotes of Leishmania mexicana to efficiently bind to membrane-bound mucin-like, O-linked glycoproteins of the sand fly Lutzomyia longipalpis, an event crucial for parasite survival, and accounts for a permissive mode of binding. Thus, direct interaction between parasite and sand fly midgut glycans are key to permitting vector competence for all forms of leishmaniasis worldwide. In addition, these studies demonstrate the feasibility of interfering with these interactions as transmission-blocking vaccines.

The secretion of extracellular polymeric substances provides an evolutionary advantage found in many organisms that can adhere to surfaces and cover themselves in a protective matrix. This ability is found in prokaryotes, archaea and eukaryotes, all of which use functionally similar polysaccharides, proteins and nucleic acids to form extracellular matrices, mucus and bioadhesive substances. These macromolecules have been investigated from the perspective of polymer biophysics, and theories to help understand adhesion, viscosity and gelling have been developed. These properties can be measured experimentally using straightforward methods such as cell counting as well as more advanced techniques such as atomic force microscopy and rheometry. An integrated understanding of the properties and uses of adhesive macromolecules across kingdoms is also important and can provide the basis for a range of biotechnological and medical applications. This article is part of the theme issue ‘Transdisciplinary approaches to the study of adhesion and adhesives in biological systems’.

Force spectroscopy was used to show that extracellular DNA (eDNA) has a pre-eminent structural role in a biofilm. The adhesive behavior of extracellular polymeric substances to poly(ethylene terephthalate), a model hydrophobic surface, was measured in response to their degradation by hydrolytic enzymes known for their biofilm dispersion potential: DNaseI, protease, cellulase, and mannanase. Only treatment with DNaseI significantly decreased the adhesive force of the model bacterium Micrococcus luteus with the surface, and furthermore this treatment almost completely eliminated any components of the biofilm maintaining the adhesion, establishing a key structural role for eDNA.

Mucus layers often provide a unique and multi-functional hydrogel interface between the epithelial cells of organisms and their external environment. Mucus has exceptional properties including elasticity, changeable rheology and an ability to self-repair by re-annealing, and is therefore an ideal medium for trapping and immobilising pathogens and serving as a barrier to microbial infection. The ability to produce a functional surface mucosa was an important evolutionary step, which evolved first in the Cnidaria, which includes corals, and the Ctenophora. This allowed the exclusion of non-commensal microbes and the subsequent development of the mucus-lined digestive cavity seen in higher metazoans. The fundamental architecture of the constituent glycoprotein mucins is also evolutionarily conserved. Although an understanding of the biochemical interactions between bacteria and the mucus layer are important to the goal of developing new antimicrobial strategies, they remain relatively poorly understood. This review summarises the physicochemical properties and evolutionary importance of mucus, which make it so successful in the prevention of bacterial infection. In addition, the strategies developed by bacteria to counteract the mucus layer are also explored.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.