The adsorption of large proteins in electrofocusing on immobilized pH gradients: I. Protein specificity and dependence on Immobiline and carrier ampholyte concentrations.
Phycoerythrin, ferritin, urease, beta-galactosidase and thyroglobulin, with molecular masses in excess of 200 kDa, adsorb and consequently fail to migrate to, and focus at, their pI positions in electrofocusing in immobilized pH gradients at a total Immobiline concentration of 20 mM while they do focus normally in pH gradients formed by carrier ampholytes. The addition of carrier ampholytes (pH range 3.5-9.5) at concentrations of 0.1 to 5% to the Immobiline-containing gels reduces adsorption (desorbs) some but not all of the 5 proteins at specific Immobiline concentrations. The adsorption is not due to water redistribution and consequent reduction in gel porosity; nor is it due to conductivity minima across the pH gradient. The hypothesis that the presence of oligomeric Immobiline contributed to the protein adsorption is the subject of the accompanying report.
Fawcett
J S
JS
Section on Macromolecular Analysis, National Institute of Child Health and Human Development, Bethesda, MD 20892.
Chrambach
A
A
D000178:Acrylamides; D000327:Adsorption; D004798:Enzymes / Q000302:isolation & purification; D005782:Gels; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing / Q000379:methods*; D008970:Molecular Weight; D011506:Proteins / Q000302:isolation & purification*
D000178:Acrylamides; D004798:Enzymes; D005782:Gels; D011506:Proteins; C059413:Immobiline