Role of EPS in mitigation of plant abiotic stress: The case of Methylobacterium extorquens PA1.

Methylobacterium extorquens is a facultative methylotrophic Gram-negative bacterium, often associated with plants, that exhibits a unique ability to grow in the presence of high methanol concentrations, which serves as a single carbon energy source. We found that M. extorquens strain PA1 secretes a mixture of different exopolysaccharides (EPSs) when grown in reference medium or in presence of methanol, that induces the secretion of a peculiar and heterogenous mixture of EPSs, with different structure, composition, repeating units, bulk and a variable degree of methylation. These factors influenced 3D structure and supramolecular assets, diffusion properties and hydrodynamic radius, and likely contribute to increase methanol tolerance and cell stability. No direct methanol involvement in the EPSs solvation shell was detected, indicating that the polymer exposure to methanol is water mediated. The presence of methanol induces no changes in size and shape of the polymer chains, highlighting how water-methanol mixtures are a good solvent for refEPS and metEPS.

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

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.

GMMA and Glycoconjugate Approaches Compared in Mice for the Development of a Vaccine against Shigella flexneri Serotype 6.

Shigella infections are one of the top causes of diarrhea throughout the world, with Shigella flexneri being predominant in developing countries. Currently, no vaccines are widely available and increasing levels of multidrug-resistance make Shigella a high priority for vaccine development. The serotype-specific O-antigen moiety of Shigella lipopolysaccharide has been recognized as a key target for protective immunity, and many O-antigen based candidate vaccines are in development. Recently, the Generalized Modules for Membrane Antigens (GMMA) technology has been proposed as an alternative approach to traditional glycoconjugate vaccines for O-antigen delivery. Here, these two technologies are compared for a vaccine against S. flexneri serotype 6. Genetic strategies for GMMA production, conjugation approaches for linkage of the O-antigen to CRM197 carrier protein, and a large panel of analytical methods for full vaccine characterization have been put in place. In a head-to-head immunogenicity study in mice, GMMA induced higher anti-O-antigen IgG than glycoconjugate administered without Alhydrogel. When formulated on Alhydrogel, GMMA and glycoconjugate elicited similar levels of persistent anti-O-antigen IgG with bactericidal activity. Glycoconjugates are a well-established bacterial vaccine approach, but can be costly, particularly when multicomponent preparations are required. With similar immunogenicity and a simpler manufacturing process, GMMA are a promising strategy for the development of a vaccine against Shigella.