Mass spectrometric analysis of the electroeluates of fluorescent proteins after preparative electrophoresis in the automated HPGE-1000 apparatus.
Bands of green fluorescent protein (GFP) and R-phycoerythrin (PHYCO) in gel electrophoresis on the automated apparatus for gel electrophoresis with periodic fluorescence scanning (HPGE), the HPGE-1000 apparatus, were retrieved from the gel by electroelution. While PHYCO was recovered in a single volume of electroeluate buffer after the predicted migration time, GFP fluorescence was lost under the same conditions and could only be recovered using multiple changes of electroeluate buffer. The multiple volumes of buffer necessitated pooling, concentration, and storage, conditions under which a minor GFP component, GFP-II, formed artifactually. PHYCO after electroelution also exhibits a minor component present in the original preparation. The electroeluate of GFP, transferred into a mass spectrometer after pooling, concentration and storage, is indistinguishable in mass from the original preparation.
Yarmola
E
E
Section on Macromolecular Analysis, Laboratory of Cellular and Molecular Biophysics, NICHD, National Institutes of Health, Bethesda, MD 20892-1580, USA.
Chrambach
A
A
Nguyen
V Q
VQ
Yergey
A L
AL
D001331:Automation; D002021:Buffers; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000032:analysis*; D013058:Mass Spectrometry / Q000379:methods*; D010799:Phycoerythrin; D011993:Recombinant Fusion Proteins / Q000032:analysis*; D013997:Time Factors
D002021:Buffers; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D010799:Phycoerythrin; D049452:Green Fluorescent Proteins