High resolution of proteins by double-inverted gradient polyacrylamide gel electrophoresis (DG-PAGE).
We have designed a new method for high resolution electrophoretic separation of proteins that have similar molecular weights. The proteins migrate first through a conventional gradient gel, in which molecular friction increases as pore size decreases. The proteins then enter an inverted sodium dodecyl sulfate (SDS) gradient gel in which friction decreases; thus, smaller molecules gradually migrate faster and achieve improved separation from larger molecules, which remain near the border between the two gels. We therefore call this technique double-inverted gradient polyacrylamide gel electrophoresis (DG-PAGE). This technique was used to resolve mixtures of aldolase, horseradish peroxidase precursors, glucose 6-phosphate dehydrogenase and pyruvate kinase. By comparison with other established methods, we show that DG-PAGE has a higher resolving power, which achieves clear separation of proteins differing as little as 0.5 kDa in molecular weight.
Zardoya
R
R
Departamento de Bioquímica y Biología Molecular IV, Universidad Complutense de Madrid, Facultad de Veterinaria, Spain.
Diez
A
A
Mason
P J
PJ
Luzzatto
L
L
Garrido-Pertierra
A
A
Bautista
J M
JM
D000818:Animals; D001709:Biotechnology; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005634:Fructose-Bisphosphate Aldolase / Q000302:isolation & purification; D005954:Glucosephosphate Dehydrogenase / Q000302:isolation & purification; D006735:Horseradish Peroxidase / Q000302:isolation & purification; D006801:Humans; D008970:Molecular Weight; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification*; D011770:Pyruvate Kinase / Q000302:isolation & purification; D011817:Rabbits; D012967:Sodium Dodecyl Sulfate
D011506:Proteins; D012967:Sodium Dodecyl Sulfate; D005954:Glucosephosphate Dehydrogenase; D006735:Horseradish Peroxidase; D011770:Pyruvate Kinase; D005634:Fructose-Bisphosphate Aldolase