Influence of the 524-VAAEIL-529 sequence of annexins A6 in their interfacial behavior and interaction with lipid monolayers.

in Journal of Colloid & Interface Science (2013), In Press

Annexin A6 (AnxA6), a calcium- and membrane-binding protein, is expressed in mammalian cells in two isoforms: AnxA6-1 and AnxA6-2, the latter lacking the 524- VAAEIL-529 sequence at the start of repeat 7 ... [more ▼]

Annexin A6 (AnxA6), a calcium- and membrane-binding protein, is expressed in mammalian cells in two isoforms: AnxA6-1 and AnxA6-2, the latter lacking the 524- VAAEIL-529 sequence at the start of repeat 7. The different intracellular localization of these two isoforms suggests distinct function in membrane dynamics. The aim of this work was to analyze the behavior of AnxA6 isoforms at the air/water interface alone and in the presence of membrane mimicking lipid monolayers. Using Langmuir technique showed that AnxA6-2 was less adsorbed to the neat air-water interface than AnxA6-1 at acidic pH and minor differences in their PM-IRRAS spectra were observed. Both isoforms exhibited similar behavior towards cholesterol monolayer. However, the interactions of AnxA6-2 with cholesterol ester monolayer were most favorable compared to AnxA6-1. Our experimental data are discussed in relation with the different intracellular localization of the two isoforms and with our constructed model of AnxA6-2 with the known crystal structure of AnxA6-1 showing the persistence of the 516-529 α- helix in AnxA6-2 despite the absence of the 524-VAAEIL-529 sequence. [less ▲]

Simulations of the Antifungal Lipopeptide Mycosubtilin in Langmuir Monolayers of Sterols

Poster (2012, July)

Interactions of the antifungal mycosubtilin with ergosterol-containing interfacial monolayers

in Biochimica et Biophysica Acta - Biomembranes (2012), 1818(5), 1302-1318

Mycosubtilin, an antimicrobial lipopeptide produced by Bacillus subtilis, is characterized by strong antifungal activities. The molecular mechanisms of its biological activities on the membranes of the ... [more ▼]

Mycosubtilin, an antimicrobial lipopeptide produced by Bacillus subtilis, is characterized by strong antifungal activities. The molecular mechanisms of its biological activities on the membranes of the sensitive yeasts or fungi have not yet been clearly elucidated. Our purpose was to mimic the mycosubtilin interactions with these membranes using various Langmuir monolayers. Since the major sterol of yeasts or fungi is ergosterol, the interactions of mycosubtilin with monolayers constituted by ergosterol, DPPC/ergosterol or DPPC/sphingomyelin/ergosterol were examined at different initial surface pressures (Πi). Plotting the mycosubtilin-induced surface pressure increases versus Πi allowed to determine that the exclusion pressures of mycosubtilin from these different monolayers is higher than the surface prevailing within the biological membranes. However, this behavior was lost when mycosubtilin was interacting with ergosteryl acetate-containing monolayers. This suggests the involvement of the sterol alcohol group in the mycosubtilin interactions within membranes. Furthermore, the behavior of mycosubtilin with stigmasterol, similar to that observed with ergosterol, differs from that previously observed with cholesterol, suggesting a role of the alkyl side chain of the sterols. The adsorption of mycosubtilin to ergosterol monolayers induced changes in the lipopeptide orientation at the air-water interface as revealed by polarization modulation infrared reflection absorption spectroscopy (PM-IRRAS). Moreover, imaging the air-water interface by Brewster angle microscopy (BAM) indicates that mycosubtilin induced changes in the organization and morphology of monolayers containing pure ergosterol with the appearance of small condensed dots, suggesting again that the target of mycosubtilin might be the ergosterol present in the membranes of the sensitive yeasts or fungi. [less ▲]

Annexins as organizers of cholesterol- and sphingomyelin-enriched membrane microdomains in Niemann-Pick type C disease.

in Cellular and Molecular Life Sciences : CMLS (2011), 69(11), 1773-1785

Growing evidence suggests that membrane microdomains enriched in cholesterol and sphingomyelin are sites for numerous cellular processes, including signaling, vesicular transport, interaction with ... [more ▼]

Growing evidence suggests that membrane microdomains enriched in cholesterol and sphingomyelin are sites for numerous cellular processes, including signaling, vesicular transport, interaction with pathogens, and viral infection, etc. Recently some members of the annexin family of conserved calcium and membrane-binding proteins have been recognized as cholesterol-interacting molecules and suggested to play a role in the formation, stabilization, and dynamics of membrane microdomains to affect membrane lateral organization and to attract other proteins and signaling molecules onto their territory. Furthermore, annexins were implicated in the interactions between cytosolic and membrane molecules, in the turnover and storage of cholesterol and in various signaling pathways. In this review, we focus on the mechanisms of interaction of annexins with lipid microdomains and the role of annexins in membrane microdomains dynamics including possible participation of the domain-associated forms of annexins in the etiology of human lysosomal storage disease called Niemann-Pick type C disease, related to the abnormal storage of cholesterol in the lysosome-like intracellular compartment. The involvement of annexins and cholesterol/sphingomyelin-enriched membrane microdomains in other pathologies including cardiac dysfunctions, neurodegenerative diseases, obesity, diabetes mellitus, and cancer is likely, but is not supported by substantial experimental observations, and therefore awaits further clarification. [less ▲]

Second harmonic generation as a tool to probe the interactions of peptides with membrane-mimicking interfacial monolayers

Conference (2011)

Interactions de la mycosubtiline avec les membranes biomimétiques. Apport de l’optique non linéaire

Scientific conference (2011)

Analyses biophysiques des interactions peptide-monocouches à l’interface air-eau

Poster (2011)

Specific interactions of mycosubtilin with cholesterol-containing artifical membranes

in Langmuir (2011), 27

Mycosubtilin is a natural antimicrobial lipopeptide produced by Bacillus subtilis strains. It is characterized by its hemolytic and strong antifungal activities. Mycosubtilin interacts with the plasma ... [more ▼]

Mycosubtilin is a natural antimicrobial lipopeptide produced by Bacillus subtilis strains. It is characterized by its hemolytic and strong antifungal activities. Mycosubtilin interacts with the plasma membranes of sensitive cells. However, the molecular mechanisms of its biological activities have not been completely elucidated. Our purpose was therefore to analyze the interactions of mycosubtilin with biological membranes by using biomimetic membranes such as Langmuir monolayers and multilayers. Structural changes of mycosubtilin, involving its peptide backbone and the side chain of its tyrosyl residue, were observed when the lipopeptide was interacting with cholesterol-containing multilayers. The interactions of mycosubtilin with monolayers constituted by pure lipids and by phosholipid/cholesterol or phospholipid/sphingomyelin/cholesterol were also examined. An original behavior of mycosubtilin toward cholesterol-containing monolayers was found. However, this original behavior was lost when mycosubtilin was interacting with pure cholesterylacetate monolayers. This suggests the involvement of the alcohol group of cholesterol in mycosubtilin-cholesterol interactions within membranes. Moreover, mycosubtilin induced changes in the organization and morphology of cholesterol-containing monolayers, and large condensed domains with different levels of condensation appeared only in the case of DPPC/sphingomyelin/cholesterol monolayer. [less ▲]

Interactions of the natural antimicrobial mycosubtilin with phospholipid membrane models

in Colloids and Surfaces B: Biointerfaces (2010), 78

Among the secondary metabolite lipopeptides produced by Bacillus subtilis, mycosubtilin is characterized by its strong antifungal activities. Even though its structure and its cellular target, the ... [more ▼]

Among the secondary metabolite lipopeptides produced by Bacillus subtilis, mycosubtilin is characterized by its strong antifungal activities. Even though its structure and its cellular target, the cytoplasmic membrane, have been determined, the molecular mechanisms of the biological activity of mycosubtilin have not been completely elucidated. In this work, the interactions between mycosubtilin and cytoplasmic membranes were modelled by using biomimetic systems such as Langmuir monolayers at the air-water interface and lipid multilamellar vesicles. The interactions of mycosubtilin with these biomimetic systems were examined, for the first time, by using specific techniques such as polarization modulation infrared reflection absorption spectroscopy, Brewster angle microscopy and high-resolution magic angle spinning NMR. Our findings indicate that mycosubtilin alone, at the air-water interface, forms a monolayer film and keeps its turn conformation. In the presence of DMPC, mycosubtilin binds to phospholipid monolayers, in a surface pressure-dependent manner. This binding results in the appearance of condensed domains which can be due to the formation of mycosubtilin clusters and/or to the lipopeptide aggregation with some phospholipid molecules and/or the formation of liquid-condensed domains of DMPC. Furthermore, in multilamellar vesicles, the mycosubtilin-DMPC interactions take place at the level of the aliphatic chains of the phospholipid because the phase transition temperature of DMPC decreased in the presence of mycosubtilin. [less ▲]

Characterization of the antimicrobial activity of mycosubtilin on the plasma membranes. A biomimetic approach

Poster (2010)

Mycosubtilin, an antimicrobial lipopeptide, is produced by Bacillus subtilis strains. It belongs to the iturin family, which is characterized by the presence of a peptide part, constituted of a constant ... [more ▼]

Mycosubtilin, an antimicrobial lipopeptide, is produced by Bacillus subtilis strains. It belongs to the iturin family, which is characterized by the presence of a peptide part, constituted of a constant chiral amino acid sequence cycled by a β-amino fatty acid (Fig. 1). As all the iturinic lipopeptides, mycosubtilin exhibits its biocide activities on cytoplasmic membrane of target cells [1]. Recently, the activity of mycosubtilin on pathogenic strains resistant to classical agents was shown [2]. However, despite many works focused on its structure and the optimization of its production, only a few studies are conducted to analyze mycosubtilin-membrane interactions. The purpose of our work was to better understand, at the molecular level, the mechanisms of the mycosubtilin activity on cytoplasmic membranes. Firstly, we modelled the mycosubtilin-membrane interactions by using biomimetic monolayers and their associated techniques (tensiometry and PM-IRRAS). After characterizing the interfacial properties of pure mycosubtilin [3], we used Langmuir films to investigate the mycosubtilin behavior when the lipopeptide reaches the external leaflet of the membrane. We found that the mycosubtilin adsorption to lipid monolayers depended on their lipid composition and the lipopeptide interaction with the membrane was facilitated by the presence of sterols. Then, we mimicked the insertion of the lipopeptide in the whole membrane by using multilamellar vesicles. FT-IR spectroscopy analysis showed that the interaction of mycosubtilin with the artificial membranes induced conformational changes of the lipopeptide only in the presence of sterol. 1. Maget-Dana R, Peypoux F. (1994) Toxicology 87:151-74. 2. Fickers P, Guez JS, Damblon C, Leclère V, Béchet M, Jacques P, Joris B. (2009) Appl. Environ. Microbiol. 75:4636-40. 3. Nasir MN, Thawani A, Kouzayha A, Besson F. (2010) Colloids Surf. B Biointerfaces 78 :17-23. [less ▲]