Two-dimensional electrophoresis with cationic detergents: a powerful tool for the proteomic analysis of myelin proteins. Part 2: analytical aspects.
The ability to analyze proteins in developing and damaged myelin will be crucial to improve our understanding of the mechanisms of myelinogenesis, dysmyelination, and demyelination. Comparative two-dimensional electrophoresis (2-DE) is a powerful approach to analyze these proteins. In part 1 of this study (see accompanying paper), a method for the 2-DE analysis of myelin proteins using the cationic detergents benzyldimethyl-n-hexadecylammonium chloride (16-BAC) and hexadecyltrimethylammonium bromide (cetyltrimethylammonium bromide; CTAB) was described. We obtained improved separation and found that 16-BAC is the most effective agent for separation in 2-DE of myelin proteins and that CTAB is the most effective agent for solubilization of myelin proteins. Here in part 2, major myelin proteins as well as membrane proteins with multitransmembrane domains were identified by mass spectrometry after 16-BAC/SDS-PAGE and CTAB/SDS-PAGE. In addition, a high-molecular-weight protein enriched in myelin fraction was identified as unconventional myosin ID using 16-BAC/SDS-PAGE, which had previously not been detected using immobilized pH gradient isoelectric focusing (IPG)/SDS-PAGE. From these results, we concluded that combinational analysis using IPG/SDS-PAGE, 16-BAC/SDS-PAGE, and CTAB/SDS-PAGE provides a powerful technique facilitating the proteomic analysis of myelin proteins in either developmental or pathological changes.
Yamaguchi
Yoshihide
Y
Department of Molecular Neurobiology, Tokyo University of Pharmacy and Life Sciences, Hachioji, Tokyo, Japan. yoshiy@ps.toyaku.ac.jp
Miyagi
Yudai
Y
Baba
Hiroko
H
D000818:Animals; D015153:Blotting, Western; D001923:Brain Chemistry; D002412:Cations; D003902:Detergents; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D008297:Male; D013058:Mass Spectrometry; D009185:Myelin Proteins / Q000737:chemistry* / Q000302:isolation & purification; D009186:Myelin Sheath / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D051381:Rats; D017208:Rats, Wistar
D002412:Cations; D003902:Detergents; D009185:Myelin Proteins