Fractional precipitation with polyethylene glycol

Polyethylene glycol (PEG) (I) is a hydrophilic polymer.  It is commercially available as preparations of different average molecular weights, the  6,000  and 20,000  materials  being most  often  used for protein isolation.

It is thought that PEG precipitates proteins by virtue  of excluding the proteins from a relatively large volume of water per PEG molecule. With an increase in PEG concentration the protein is thus eventually brought to  its  solubility limit.  It  has  been empirically  observed  that,  at equimolar concentrations,  large  proteins  are  precipitated  at  lower  PEG concentrations than small proteins.  For this  reason  PEG precipitation  is especially suited to the isolation of large proteins and even particles,  such as viruses.  PEG-precipitation may  thus  be  thought  of  as  “molecular exclusion precipitation”.

The effect of protein concentration has not been explored in detail but it would appear  from  limited  data that  in  PEG precipitation  proteins behave much as type I proteins  do in salting out,  i.e. the  higher the protein concentration, the  less precipitant  is required to initiate precipitation.

After precipitation,  it is difficult to  separate  the  PEG from  the protein. PEG is acetone soluble, however, and acetone precipitation of the protein can be used to  separate  it  from  the  PEG.  It  may not  always be necessary to remove the PEG since many of the subsequent uses of the protein may be tolerant of the presence of PEG.  It should be noted, though, that  PEG can alter the  behaviour of proteins  during molecular exclusion chromatography.

A practical  difficulty  in  the  use of PEG is that  it  absorbs UV light at 280 nm, and also interferes  with the  Lowry protein  assay.  Protein  can, however, be measured in the presence of PEG using the biuret assay.

References

Dennison, C. (2002). A guide to protein isolation . School of Molecular mid Cellular Biosciences, University of Natal . Kluwer Academic Publishers new york, Boston, Dordrecht, London, Moscow .