Exploring the Molecular Interactions of Symmetrical and Unsymmetrical Selenoglycosides with Human Galectin-1 and Galectin-3

Abstract

Galectins (Gals) are small cytosolic proteins that bind β-galactoside residues via their evolutionarily conserved carbohydrate recognition domain. Their dysregulation has been shown to be associated with many diseases. Consequently, targeting galectins for clinical applications has become increasingly relevant to develop tailored inhibitors selectively for one galectin. Accordingly, binding studies providing the molecular details of the interaction between galectin and inhibitor may be useful for the rational design of potent and selective antagonists. Gal-1 and Gal-3 are among the best-studied galectins, mainly for their roles in cancer progression; therefore, the molecular details of their interaction with inhibitors are demanded. This work gains more value by focusing on the interaction between Gal-1 and Gal-3 with the selenylated analogue of the Gal inhibitor thiodigalactose, characterized by a selenoglycoside bond (SeDG), and with unsymmetrical diglycosyl selenides (unsym(Se). Gal-1 and Gal-3 were produced heterologously and biophysically characterized. Interaction studies were performed by ITC, NMR spectroscopy, and MD simulation, and thermodynamic values were discussed and integrated with spectroscopic and computational results. The 3D complexes involving SeDG when interacting with Gal-1 and Gal-3 were depicted. Overall, the collected results will help identify hot spots for the design of new, better performing, and more specific Gal inhibitors.

Directing in Vitro Selection towards G‐quadruplex‐forming Aptamers to Inhibit HMGB1 Pathological Activity

“…With the aim of validating the ELONA assay and prove the specificity of the selected sequences for HMGB1, the described assay was also used to evaluate the affinity of the best‐performing aptamers for the following additional targets of biological interest: i) VEGF‐A, which is the protein used for the negative selection step in the SELEX procedure, in consideration of its high serum levels in cancer and inflammation, [42,43] analogously to HMGB1; ii) Galectin‐3, [56] which is a circulating protein involved in several biological processes, including apoptosis, cell proliferation and inflammation; iii) Siglec‐7, [57] a membrane protein found in innate lymphoid natural killer (NK) cells, blood monocytes and dendritic cells, involved in the maintenance of immune homeostasis; iv) CD19, [58] a membrane protein expressed on the surface of B and plasma cells involved in establishing intrinsic B cell signaling thresholds. In addition, VEGF‐A is also a G4 binding protein (G4BP) which showed high affinity for parallel G‐quadruplex aptamers [43,51,52,59] .…”

Directing in Vitro Selection towards G‐quadruplex‐forming Aptamers to Inhibit HMGB1 Pathological Activity

“…With the aim of validating the ELONA assay and prove the specificity of the selected sequences for HMGB1, the described assay was also used to evaluate the affinity of the best‐performing aptamers for the following additional targets of biological interest: i) VEGF‐A, which is the protein used for the negative selection step in the SELEX procedure, in consideration of its high serum levels in cancer and inflammation, [42,43] analogously to HMGB1; ii) Galectin‐3, [56] which is a circulating protein involved in several biological processes, including apoptosis, cell proliferation and inflammation; iii) Siglec‐7, [57] a membrane protein found in innate lymphoid natural killer (NK) cells, blood monocytes and dendritic cells, involved in the maintenance of immune homeostasis; iv) CD19, [58] a membrane protein expressed on the surface of B and plasma cells involved in establishing intrinsic B cell signaling thresholds. In addition, VEGF‐A is also a G4 binding protein (G4BP) which showed high affinity for parallel G‐quadruplex aptamers [43,51,52,59] .…”

Biophysical and Structural Characterization of the Interaction between Human Galectin-3 and the Lipopolysaccharide from Pseudomonas aeruginosa

“…As already described [ 9 ], the spectrum of Gal3 is not a typical spectrum of a protein with beta-sheets but displays particular characteristics in its topological arrangement, such as the length of the filaments, intra/intersheet twists and β-turns producing a spectrum with a minimum of around 220 nm. The far-UV Gal3 CRD spectrum registered in the presence of an increasing concentration of LPS pa showed the partial denaturation of Gal3 CRD , as corroborated by the disappearance of the positivity at around 200 nm.…”

“…The titration experiments showed a decrease in fluorescence emission as a function of LPSpa concentration, demonstrating that interaction with LPSpa truly takes place and suggesting the occurrence of an LPSpa-induced conformational change toward a more compact structure. Moreover, since no λ shift can be observed in the fluorescence spectra upon As already described [9], the spectrum of Gal3 is not a typical spectrum of a protein with beta-sheets but displays particular characteristics in its topological arrangement, such as the length of the filaments, intra/intersheet twists and β-turns producing a spectrum with a minimum of around 220 nm. The far-UV Gal3 CRD spectrum registered in the presence of an increasing concentration of LPSpa showed the partial denaturation of Gal3 CRD , as corroborated by the disappearance of the positivity at around 200 nm.…”

“…These proteins play an important role in several biological processes [ 7 ] and are therefore becoming emerging targets for diagnostic and therapeutic approaches, and several inhibitors, mainly of a glycosidic nature, have been identified and characterised [ 8 , 9 , 10 , 11 , 12 , 13 ].…”