Overexpressed genetically modified proteins are used for a multitude of industrial, pharmaceutical, and agricultural applications. Insoluble and misfolded proteins are a common problem during the production. Efficiently recovering the correctly folded form is a considerable challenge.
Gregory A. Weiss and co-workers, University of California, Irvine, CA, USA, have sucessfully refolded lysozyme from boiled hen egg whites, demonstrating the efficiency of their method of protein recovery. The researchers use shear stress within thin fluid films in a ‘vortex fluid device’ (VFD), a rapidly spinning glass tube. In this method, a glass cylinder (10 mm by 16 cm) is spun rapidly (5 krpm) at a 45° angle. At high rotational speeds, the solution within the sample tube forms micrometer-thick, thin fluid films, which flow with the same speed and direction as the wall of the glass tube. The rotating glass tube generates a velocity gradient within the thin fluid film, which introduces shear stress into the solution.
The team recovered active lysozyme from previously cooked and thus denatured hen egg whites, as demonstrated by an activity assay, after spinning times of only 5 minutes. After VFD processing, the Circular dichroism (CD) of identical hen egg white lysozyme (HEWL) samples demonstrated partial recovery of secondary structure compared to the native lysozyme. The refolding of HEWL from the complex mixture of boiled egg whites appeared less efficient than recovery of the folded protein isolated from inclusion bodies. In egg whites, the mechanical energy of the VFD could be misdirected to the other >96 % of proteins present.
Until now, valuable proteins have to be produced in optimized mammalian cell lines, where misfolding is not a common issue, or recovered by laborious processing of aggregated protein after expression in yeast or Escherichia coli. Compared to overnight dialysis, conventionally used for the latter, the reported method is remarkably fast. This approach could dramatically reduce costs for cancer treatments, food production and other segments of the biotechnology industry.
- Shear-Stress-Mediated Refolding of Proteins from Aggregates and Inclusion Bodies,
Tom Z. Yuan, Callum F. G. Ormonde, Stephan T. Kudlacek, Sameeran Kunche, Joshua N. Smith, William A. Brown, Kaitlin M. Pugliese, Tivoli J. Olsen, Mariam Iftikhar, Colin L. Raston, Gregory A. Weiss,
Also of Interest
- How to Launch a Hen Suddenly into Near-earth Orbit,
ChemViews Mag. 2012.
- Boiled Eggs: Soft and Hard — Part 1,
ChemViews Mag. 2012.