Browsing by Subject "Crosslinking"

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    Detergent-labile, supramolecular assemblies of KcsA: relative abundance and interactions involved
    (Elsevier, 2012-09-27) Giudici Besseghini, Ana Marcela; Molina Gallego, María Luisa; Ayala Torres, José Leonardo; Montoya Díaz, Estefanía; Renart Pérez, María Lourdes; Fernández Carvajal, Asia María; Encinar Hidalgo, José Antonio; Ferrer Montiel, Antonio Vicente; Poveda Larrosa, José Antonio; González Ros, José Manuel; Bioquímica y Biología Molecular B e Inmunología
    In this work, we illustrate the ability of the prokaryotic potassium channel KcsA to assemble into a variety of supramolecular clusters of defined sizes containing the tetrameric KcsA as the repeating unit. Such clusters, particularly the larger ones, are markedly detergent-labile and thus, disassemble readily upon exposure to the detergents commonly used in protein purification or conventional electrophoresis analysis. This is a reversible process, as cluster re-assembly occurs upon detergent removal and without the need of added membrane lipids. Interestingly, the dimeric ensemble between two tetrameric KcsA molecules are quite resistant to detergent disassembly to individual KcsA tetramers and along with the latter, are likely the basic building blocks through which the larger clusters are organized. As to the proteins domains involved in clustering, we have observed disassembly of KcsA clusters by SDS-like alkyl sulfates. As these amphiphiles bind to inter-subunit, “non-annular” sites on the protein, these observations suggest that such sites also mediate channel–channel interactions leading to cluster assembly.
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    Effect of chemical crosslinking on the swelling and shrinking properties of thermal and pH-responsive Chitosan hydrogels
    (Wiley, 2003-10-07) Heras, Ángeles; Aranaz, Inmaculada; Galed, Gemma; Fernández Valle, María E.; Argüelles Monal, Waldo; Goycoolea Valencia, Francisco Martín; Biología Celular e Histología
    The ability to form a gel through the physical or chemical crosslinking of chitosan has been well documented. In an attempt to mimic biological systems, thermal and pH-sensitive chitosan cylindrical hydrogels were produced by a combination of physical and chemical crosslinking processes. To this end, chitosan hydrogels prepared from alkali chitin were molded in cylinders and, once washed, were further crosslinked with glutaraldehyde at stoichiometric ratios, R (-[–CH=O]/[–NH2]), of 1.61 and 3.22 x10^-2. Variation in swelling as a result of stepwise changes in temperature between 40 and 2 °C at pH values of 7.0, 7.6, and 8.0 revealed that the system responds in markedly different manners dependent upon the pH. At pH 7.0, cooling from 40 to 2 °C results in contraction of the gel network structure. While raising the temperature from 2 to 40 °C leads to a rapid swelling response (i.e., ca. a twofold increase in the amount of solvent uptake). Subsequent cooling to 2 °C is accompanied by a new contraction cycle. At pH 7.6 the temperature dependence of the swelling–contraction behavior is exactly the opposite of that observed at pH 7.0.Very similar trends were observed for the gels at both degrees of crosslinking. The swelling–shrinking behavior observed in gels of pH 7.6, is similar in kind to that of uncrosslinked gels and is interpreted in terms of a lower critical solution temperature (LCST) volume phase transition, driven by hydrophobic association, presumably involving residual acetyl groups in the chitin. The results at pH 7.0 suggest that the slight ionization of the –NH3+ groups leads to the destruction of the hydrophobic hydration thus effectively reversing the negative thermal shrinking.