Two-dimensional gel electrophoresis of wool intermediate filament proteins.
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) methods have been used to provide high-resolution separation of wool intermediate filament proteins (IFPs). An improved method of extraction was developed based on a previously published method. The improved method for extraction eliminates the use of dialysis and freeze-drying between the extraction and rehydration steps, allowing the extraction and rehydration for the first dimension gel to be achieved in one day. Improvements to the method for maintaining reducing conditions and chaotrope constitution, combined with low %T polyacrylamide gels, allowed the high-resolution separation of the two keratin IFP families and their individual family members. The IFPs were separated to produce a clearly defined spot pattern of higher intensity, with numerous minor spots not previously observed, and a marked improvement in the vertical resolution. Further work to analyse the composition of each of the protein spots has been made much easier by being able to separate the IFPs into discrete spots.
Paton
Louise N
LN
School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand University of Canterbury, New Zealand.
Gerrard
Juliet A
JA
Bryson
Warren G
WG
D000818:Animals; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D007381:Intermediate Filament Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D007526:Isoelectric Point; D010084:Oxidation-Reduction; D012756:Sheep; D014935:Wool / Q000737:chemistry* / Q000378:metabolism*
D007381:Intermediate Filament Proteins