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Pathway for unfolding of ubiquitin in sodium dodecyl sulfate, studied by capillary electrophoresis.

ABSTRACT: 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.

SUBMITTER: Schneider GF

PROVIDER: S-EPMC2688835 | biostudies-literature | 2008 Dec

REPOSITORIES: biostudies-literature

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Pathway for unfolding of ubiquitin in sodium dodecyl sulfate, studied by capillary electrophoresis.

Schneider Grégory F GF Shaw Bryan F BF Lee Andrew A Carillho Emanuel E Whitesides George M GM

Journal of the American Chemical Society 20081201 51

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 complex ...[more]

PMID: 19035631

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Project description:The number and type of synthetic chemicals that are being produced worldwide continues to increase significantly. While these industrial chemicals provide numerous benefits, there is no doubt that some have potential to damage the environment and health. Toxicity must be evaluated and use must be carefully controlled and monitored in order to minimize potential damage. DNA microarray technology has become an important new technique in toxicology. We are using the yeast Saccharomyces cerevisiae as a model organism for toxicological study because it is a simple, fast-growing eukaryote that has been thoroughly characterized. In order to evaluate toxicity by newly synthesized or mixture chemicals, toxicity-induced gene expression alteration profiles by known chemicals should be collected. In our study, cells need to be exposed with same experimental cellular condition, semi lethal (IC50), respectively. In the case of SDS (CAS; 151-21-3), the exposure dose was decided as 0.01% by growth curve with continuously diluted exposure. SDS is an anionic surfactant that is suspected to be gastrointestinal or liver toxicant. // Toxicity of anionic detergents determined by Saccharomyces cerevisiae microarray analysis: Sodium n-dodecyl benzene sulfonate (LAS) and sodium dodecyl sulfate (SDS) are popular anionic detergents (surfactants) that are used worldwide and the toxicities of these chemicals have been characterized. We applied these chemicals in a DNA microarray bioassay and determined that the microarray data reflects previous findings and also provides some new information about anionic detergent toxicity. The mRNA expression profiles suggest that LAS and SDS cause damage to membranes and alterations in carbon metabolism, and induce the oxidative stress response. We also found that LAS and SDS induce the pleiotropic drug-resistance network, and that LAS and SDS may be pumped out of yeast cells by this network. Hierarchical clustering of the expression profiles showed that LAS and SDS cause similar features of toxicity and that the toxicity is similar to that of capsaicin but different from that of cadmium and mercury. Keywords: stress response

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Pathway for unfolding of ubiquitin in sodium dodecyl sulfate, studied by capillary electrophoresis.

Publications

Pathway for unfolding of ubiquitin in sodium dodecyl sulfate, studied by capillary electrophoresis.

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