Isoelectric focusing of sparingly soluble proteins in immobilized pH gradients, exemplified by microvillar membrane hydrolases.
A novel fractionation technique is described for analysis of membrane-bound enzymes and sparingly soluble proteins: isoelectric focusing in a mixed-type matrix, containing a primary, immobilized pH gradient with a superimposed, secondary carrier ampholyte pH gradient. Three microvilli hydrolases: dipeptidyl peptidase IV, gamma-glutamyl transferase and alkaline phosphatase exhibit an array of sharply focused, enzyme active bands in the pH 4-6.5 range. The separation pattern obtained is by far superior to any separation achieved by either technique separately.
D000818:Animals; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000032:analysis*; D007525:Isoelectric Focusing / Q000379:methods*; D007668:Kidney / Q000201:enzymology; D008297:Male; D008871:Microvilli / Q000201:enzymology*; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility; D013379:Substrate Specificity
D006867:Hydrolases