Pathway for unfolding of ubiquitin in sodium dodecyl sulfate, studied by capillary electrophoresis.
This paper characterizes the complexes formed by a small protein, ubiquitin (UBI), and a negatively charged surfactant, sodium dodecyl sulfate (SDS), using capillary electrophoresis (CE), circular dichroism (CD), and amide hydrogen-deuterium exchange (HDX; as monitored by mass spectroscopy, MS). Capillary electrophoresis of complexes of UBI and SDS, at apparent equilibrium, at concentrations of SDS ranging from sub-micellar and sub-denaturing to micellar and denaturing, revealed multiple complexes of UBI and SDS of the general composition UBI-SDS(n). Examination of electrophoretic mobilities of complexes of UBI and SDS as a function of the concentration of SDS provided a new way to characterize the interaction of this protein with SDS and established key characteristics of this system: e.g., the reversibility of the formation of the complexes, their approximate chemical compositions, and the pathway of SDS binding to UBI. The work identified, in addition to SDS-saturated UBI, at least six groups of complexes of UBI with SDS, within which four groups were populated with complexes of distinct stoichiometries: UBI-SDS(approximately 11), UBI-SDS(approximately 25), UBI-SDS(approximately 33), and UBI-SDS(approximately 42). CD spectroscopy and amide HDX of the UBI-SDS(n) complexes suggested that many of the UBI-SDS(n) complexes (n > 11) have greater alpha-helical content than native UBI. Capillary electrophoresis provides a level of detail about interactions of proteins and SDS that has not previously been accessible, and CE is an analytical and biophysical method for studies of interactions of proteins and surfactants that is both convenient and practical. This study sheds light on the formation of the enigmatic protein-SDS complexes formed during SDS polyacrylamide gel electrophoresis and brings a new tool to the study of proteins and detergents.
Schneider
Grégory F
GF
Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts 02138, USA.
Shaw
Bryan F
BF
Lee
Andrew
A
Carillho
Emanuel
E
Whitesides
George M
GM
D000577:Amides / Q000737:chemistry; D000818:Animals; D001671:Biochemistry / Q000379:methods; D002417:Cattle; D002942:Circular Dichroism; D019075:Electrophoresis, Capillary / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008055:Lipids / Q000737:chemistry; D008823:Micelles; D008968:Molecular Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry*; D025801:Ubiquitin / Q000737:chemistry*
D000577:Amides; D008055:Lipids; D008823:Micelles; D025801:Ubiquitin; D012967:Sodium Dodecyl Sulfate