Dataset Information

Influence of fluorocarbon and hydrocarbon acyl groups at the surface of bovine carbonic anhydrase II on the kinetics of denaturation by sodium dodecyl sulfate.

ABSTRACT: This paper examines the influence of acylation of the Lys-?-NH(3)(+) groups of bovine carbonic anhydrase (BCA, EC 4.2.1.1) to Lys-?-NHCOR (R = -CH(3), -CH(2)CH(3), and -CH(CH(3))(2), -CF(3)) on the rate of denaturation of this protein in buffer containing sodium dodecyl sulfate (SDS). Analysis of the rates suggested separate effects due to electrostatic charge and hydrophobic interactions. Rates of denaturation (k(Ac,n)) of each series of acylated derivatives depended on the number of acylations (n). Plots of log k(Ac,n) vs n followed U-shaped curves. Within each series of derivatives, rates of denaturation decreased as n increased to ?7; this decrease was compatible with increasingly unfavorable electrostatic interactions between SDS and protein. In this range of n, rates of denaturation also depended on the choice of the acyl group as n increased to ?7, in a manner compatible with favorable hydrophobic interactions between SDS and the -NHCOR groups. As n increased in the range 7 < n < 14, however, rates of denaturation stayed approximately constant; analysis suggested that these rates were compatible with an increasingly important contribution to denaturation that depended both on the net negative charge of the protein and on the hydrophobicity of the R group. The mechanism of denaturation thus seems to change with the extent of acylation of the protein. For derivatives with the same net electrostatic charge, rates of denaturation increased with the acyl group (by a factor of ?3 for n ? 14) in the order CH(3)CONH- < CH(3)CH(2)CONH- < (CH(3))(2)CHCONH- < CF(3)CONH-. These results suggested that the hydrophobicity of CF(3)CONH- is slightly greater (by a factor of <2) than that of RHCONH- with similar surface area.

SUBMITTER: Lee A

PROVIDER: S-EPMC3034637 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Influence of fluorocarbon and hydrocarbon acyl groups at the surface of bovine carbonic anhydrase II on the kinetics of denaturation by sodium dodecyl sulfate.

Lee Andrew A Mirica Katherine A KA Whitesides George M GM

The journal of physical chemistry. B 20101223 5

This paper examines the influence of acylation of the Lys-ε-NH(3)(+) groups of bovine carbonic anhydrase (BCA, EC 4.2.1.1) to Lys-ε-NHCOR (R = -CH(3), -CH(2)CH(3), and -CH(CH(3))(2), -CF(3)) on the rate of denaturation of this protein in buffer containing sodium dodecyl sulfate (SDS). Analysis of the rates suggested separate effects due to electrostatic charge and hydrophobic interactions. Rates of denaturation (k(Ac,n)) of each series of acylated derivatives depended on the number of acylations ...[more]

PMID: 21182314

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Project description:This article describes the use of capillary electrophoresis (CE) to examine the influence of different cations (C(+); C(+) = Na(+) and tetra-n-alkylammonium, NR(4)(+), where R = Me, Et, Pr, and Bu) on the rates of denaturation of bovine carbonic anhydrase II (BCA) in the presence of anionic surfactant dodecylsulfate (DS(-)). An analysis of the denaturation of BCA in solutions of Na(+)DS(-) and NR(4)(+)DS(-) (in Tris-Gly buffer) indicated that the rates of formation of complexes of denatured BCA with DS(-) (BCA(D)-DS(-)(n,sat)) are indistinguishable and independent of the cation below the critical micellar concentration (cmc) and independent of the total concentration of DS(-) above the cmc. At concentrations of C(+)DS(-) above the cmc, BCA denatured at rates that depended on the cation; the rates decreased by a factor >10(4) in the order of Na(+) ≈ NMe(4)(+) > NEt(4)(+) > NPr(4)(+) > NBu(4)(+), which is the same order as the values of the cmc (which decrease from 4.0 mM for Na(+)DS(-) to 0.9 mM for NBu(4)(+)DS(-) in Tris-Gly buffer). The relationship between the cmc values and the rates of formation of BCA(D)-DS(-)(n,sat()) suggested that the kinetics of denaturation of BCA involve the association of this protein with monomeric DS(-) rather than with micelles of (C(+)DS(-))(n). A less-detailed survey of seven other proteins (α-lactalbumin, β-lactoglobulin A, β-lactoglobulin B, carboxypeptidase B, creatine phosphokinase, myoglobin, and ubiquitin) showed that the difference between Na(+)DS(-) and NR(4)(+)DS(-) observed with BCA was not general. Instead, the influence of NR(4)(+) on the association of DS(-) with these proteins depended on the protein. The selection of the cation contributed to the properties (including the composition, electrophoretic mobility, and partitioning behavior in aqueous two-phase systems) of aggregates of denatured protein and DS(-). These results suggest that the variation in the behavior of NR(4)(+)DS(-) with changes in R may be exploited in methods used to analyze and separate mixtures of proteins.

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

Dataset Information

Influence of fluorocarbon and hydrocarbon acyl groups at the surface of bovine carbonic anhydrase II on the kinetics of denaturation by sodium dodecyl sulfate.

Publications

Influence of fluorocarbon and hydrocarbon acyl groups at the surface of bovine carbonic anhydrase II on the kinetics of denaturation by sodium dodecyl sulfate.

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