Conversion of a transmembrane to a water-soluble protein complex by a single point mutation
Proteins exist in one of two generally incompatible states: either membrane associated or soluble. Pore-forming proteins are exceptional because they are synthesized as a water-soluble molecule but end up being located in the membrane -- that is, they are nonconstitutive membrane proteins. Here we report the pronounced effect of the single point mutation Y221G of the pore-forming toxin aerolysin. This mutation blocks the hemolytic activity of the toxin but does not affect its initial structure, its ability to bind to cell-surface receptors or its capacity to form heptamers, which constitute the channel-forming unit. The overall structure of the Y221G protein as analyzed by cryo-negative staining EM and three-dimensional reconstruction is remarkably similar to that of the wild type heptamer. The mutant protein forms a mushroom-shaped complex whose stem domain is thought to be within the membrane in the wild type toxin. In contrast to the wild type heptamer, which is a hydrophobic complex, the Y221G heptamer is fully hydrophilic. This point mutation has, therefore, converted a normally membrane-embedded toxin into a soluble complex.
Keywords: Aeromonas hydrophila ; Bacterial Toxins/chemistry/*genetics/metabolism ; Membrane Proteins/chemistry/genetics/metabolism ; *Point Mutation ; Pore Forming Cytotoxic Proteins ; Protein Structure ; Quaternary ; Protein Structure ; Tertiary ; Solubility
Department of Genetics and Microbiology, University of Geneva, 1 rue Michel Servet, 1211 Geneva, Switzerland.
Record created on 2009-01-30, modified on 2016-08-08