Unfolding Individual Als5p Adhesion Proteins on Live Cells.

Elucidating the molecular mechanisms behind the strength and mechanics of cell adhesion proteins is of central importance in cell biology and offers exciting avenues for the identification of potential drug targets. Here we use single-molecule force spectroscopy to investigate the adhesive and mechanical properties of the widely expressed Als5p cell adhesion protein from the opportunistic pathogen Candida albicans . We show that the forces required to unfold individual tandem repeats of the protein are in the 150-250 pN range, both on isolated molecules and on live cells. We also find that the unfolding probability increases with the number of tandem repeats and correlates with the level of cell adherence. We suggest that the modular and flexible nature of Als5p conveys both strength and toughness to the protein, making it ideally suited for cell adhesion. The single-molecule measurements presented here open new avenues for understanding the mechanical properties of adhesion molecules from mammalian and microbial cells and may help us to elucidate their potential implications in diseases such as inflammation, cancer, and infection

Copper oxide in layered tin oxide with intracrystalline microporosity for oxidative imination of toluene and p-xylene

Homoepitaxial layered structure of tin oxide, SnO2 containing low amount of copper oxide, CuO (named as CuOSnO2) was found to convert toluene and p-xylene into value-added compounds. The CuO-SnO2 with microporous channels and expanded lattice planes allowed the high diffusion of toluene and p-xylene. Such characteristics features made the oxidative coupling of toluene and p-xylene with various amines resulting in different imines with good yield (70-92%) within 12 h. The reaction proceeded with the selective transformation from toluene to benzaldehyde and its condensation with amines at 120degree celsius in presence of H2O2 as oxidant. Theoretical investigations were done to understand to know the thermodynamic feasibility of the reaction. The same study also suggested the involvement of weak supramolecular forces like cation (Sn4+)-pi interactions in stabilizing the intermediate state during the course of the reaction. The thermally stable CuO-SnO2 catalysts with moderate to strong basic sites appeared as a suitable recyclable catalyst for the oxidative imination raction

Global Cell Surface Conformational Shift Mediated by a Candida albicans Adhesin

Candida albicans maintains both commensal and pathogenic states in humans. Both states are dependent on cell surface-expressed adhesins, including those of the Als family. Heterologous expression of Als5p at the surface of Saccharomyces cerevisiae results in Als5p-mediated adhesion to various ligands, followed by formation of multicellular aggregates. Following adhesion of one region of the cell to fibronectin-coated beads, the entire surface of the cells became competent to mediate cell-cell aggregation. Aggregates formed in the presence of metabolic inhibitors or signal transduction inhibitors but were reduced in the presence of 8-anilino-1-naphthalene-sulfonic acid (ANS) or Congo Red (CR), perturbants that inhibit protein structural transitions. These perturbants also inhibited aggregation of C. albicans. An increase in ANS fluorescence, which accompanied Als-dependent cellular adhesion, indicated an increase in cell surface hydrophobicity. In addition, C. albicans and Als5p-expressing S. cerevisiae showed an aggregation-induced birefringence indicative of order on the cell surface. The increase in birefringence did not occur in the presence of the aggregation disruptants ANS and CR. These results suggest a model for Als5p-mediated aggregation in which an adhesion-triggered change in the conformation of Als5p propagates around the cell surface, forming ordered aggregation-competent regions

Threonine-Rich Repeats Increase Fibronectin Binding in the Candida albicans Adhesin Als5p

Commensal and pathogenic states of Candida albicans depend on cell surface-expressed adhesins, including those of the Als family. Mature Als proteins consist of a 300-residue N-terminal region predicted to have an immunoglobulin (Ig)-like fold, a 104-residue conserved Thr-rich region (T), a central domain of a variable number of tandem repeats (TR) of a 36-residue Thr-rich sequence, and a heavily glycosylated C-terminal Ser/Thr-rich stalk region, also of variable length (N. K. Gaur and S. A. Klotz, Infect. Immun. 65: 5289-5294, 1997). Domain deletions in ALS5 were expressed in Saccharomyces cerevisiae to excrete soluble protein and for surface display. Far UV circular dichroism indicated that soluble Ig-T showed a single negative peak at 212 nm, consistent with previous data indicating that this region has high β-sheet content with very little α-helix. A truncation of Als5p with six tandem repeats (Ig-T-TR(6)) gave spectra with additional negative ellipticity at 200 nm and, at 227 to 240 nm, spectra characteristic of a structure with a similar fraction of β-sheet but with additional structural elements as well. Soluble Als5p Ig-T and Ig-T-TR(6) fragments bound to fibronectin in vitro, but the inclusion of the TR region substantially increased affinity. Cellular adhesion assays with S. cerevisiae showed that the Ig-T domain mediated adherence to fibronectin and that TR repeats greatly increased cell-to-cell aggregation. Thus, the TR region of Als5p modulated the structure of the Ig-T region, augmented cell adhesion activity through increased binding to mammalian ligands, and simultaneously promoted fungal cell-cell interactions