Effect of temperature and low pH on structure and stability of matrix porin in micellar detergent solutions.
Thermal and low pH stabilities of matrix porin (Omp F) solubilized in the micellar solutions of ionic (SDS) and nonionic detergents were investigated by the methods of circular dichroism, intrinsic fluorescence, light scattering and sedimentation velocity. The stability of porin structure in solution is much higher in the presence of beta-octyl glucoside than with SDS. In the presence of SDS, sharp transitions were detected by all parameters measured, above 55 degrees C at neutral pH and below pH 4.5 at 20 degrees C. These transitions involve at least three concomitant processes: unfolding of protein, dissociation of trimers to monomers and the disruption of the protein-detergent micellar complexes, all events being irreversible in the presence of SDS. The nonionic detergent, beta-octyl glucoside, increases the stability of porin in acidic conditions, since neither dissociation nor denaturation was observed in the pH region between 7.5 and 2.0. However, at pH less than 3.5, small, reversible changes in protein structure became evident. The thermal stability of porin is also increased by beta-octyl glucoside as evidenced by a transition temperature 15-20 degrees C higher as compared to SDS. A considerable degree of native porin structure was regained after heat treatment in the presence of beta-octyl glucoside, though the reconstituted trimers were not identical to the native ones. The addition of lipopolysaccharide and divalent cations (Ca2+, Mg2+) to the experimental system did not improve the thermal reversibility.
D001425:Bacterial Outer Membrane Proteins; D002942:Circular Dichroism; D003460:Crystallization; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D006863:Hydrogen-Ion Concentration; D046911:Macromolecular Substances; D008823:Micelles; D018272:Porins; D011485:Protein Binding; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013696:Temperature
D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D046911:Macromolecular Substances; D008823:Micelles; D018272:Porins