Solubilization and stabilization of bacteriophage MS2 in organic solvents.
Several techniques were examined for the solubilization of bacteriophage MS2 in organic solvents. Direct extraction of the MS2 from an aqueous phase into isooctane containing 2 mM AOT, a proven approach for the organic solubilization of many proteins, was not successful. However, predried samples of MS2 were solubilized through the direct addition of organic solvents containing 500 mM AOT. As an alternative procedure, reverse micelles containing aqueous solutions of MS2 were prepared in isooctane using AOT, dehydrated through solvent evaporation and azeotropic drying, and resolubilized in a solvent of choice. The structure and microenvironment of organic-solubilized MS2 were investigated by UV absorbance, the fluorescence emission of an attached solvatochromatic dye, tryptophan fluorescence, and atomic force microscopy, all of which contributed evidence for a fully assembled capsid in the organic solvent. The solubilized MS2 was derivatized with stearic acid in chloroform, illustrating that bioconjugation reactions can be performed on organic-solubilized capsids using reagents that are completely insoluble in water. Furthermore, the organic-solubilized phage remained infectious after heating at 90 degrees C for 20 min, whereas phage in aqueous buffer or dried with nitrogen were nonviable following the heat treatment protocol. The extended range of available chemical modifications and the enhanced thermal stability of the organic-solubilized capsids bodes well for the formulation of storage-stable vaccines predicated on reactions in or exposure to organic media.
Johnson
Harvey R
HR
Department of Chemical Engineering, University of California, Berkeley, California 94720, USA.
Hooker
Jacob M
JM
Francis
Matthew B
MB
Clark
Douglas S
DS
D001709:Biotechnology; D002213:Capsid / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000502:physiology; D005822:Genetic Vectors; D006358:Hot Temperature; D017909:Levivirus / Q000737:chemistry / Q000378:metabolism*; D008823:Micelles; D011489:Protein Denaturation; D012995:Solubility; D012997:Solvents; D014764:Viral Proteins / Q000737:chemistry*
D008823:Micelles; D012997:Solvents; D014764:Viral Proteins