Highly basic myelin and oligodendrocyte proteins analyzed by NEPHGE-two-dimensional gel electrophoresis: Recognition of novel developmentally regulated proteins
Y. Yamaguchi et Se. Pfeiffer, Highly basic myelin and oligodendrocyte proteins analyzed by NEPHGE-two-dimensional gel electrophoresis: Recognition of novel developmentally regulated proteins, J NEUROSC R, 56(2), 1999, pp. 199-205
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) provides high
resolution separation of proteins and offers a powerful method for their id
entification and characterization. Since many myelin-specific proteins are
highly basic, they cannot readily be analyzed by standard isoelectric focus
ing (IEF)2D-PAGE that affords separation primarily in the isoelectric point
s (pI) range of 4-8. An alternative method, nonequilibrium pH gradient elec
trophoresis (NEPHGE)-2D-PAGE, can provide excellent resolution of highly ba
sic proteins. In the present study, we have optimized the NEPHGE-2D-PAGE pr
otocol for the analysis of myelin proteins with basic pIs, and provide a NE
PHGE-2D-PAGE map based on size, pI, and immunoreactivity (Western blot) of
myelin basic protein (MBP), 2',3'-cyclic-nucleotide 3'-phosphodiesterase (C
NP), myelin proteolipid protein (PLP), and its smaller spliced variant DM20
, myelin/oligodendrocyte glycoprotein (MOG) and oligodendrocyte-specific pr
otein (OSP), We have also demonstrated, by analyzing metabolically radiolab
eled oligodendrocytes in culture at specific stages of the developmental li
neage, the developmentally up-regulated expressions of several undefined, o
ligodendrocyte, basic membrane proteins during oligodendrocyte differentiat
ion. We suggest that this approach offers an important tool for identifying
and characterizing the plethora of uncharacterized myelin proteins. J. Neu
rosci. Res. 56:199-205, 1999, (C) 1999 Wiley-Liss, Inc.