Abstract
Critical micelle concentrations (CMCs) of palmitoyl-CoA were determined by surface tension, conductivity, and fluorimetric measurements in a variety of buffers at several pH values and ionic strengths. They ranged from 7 to 250 microM and were frequently an order of magnitude higher than most reported values. The CMCs of stearoyl-CoA and oleoyl-CoA, determined fluorimetrically, were also high and consistent with the expected effects of chain length and unsaturation. The effects of ionic strength and temperature were analyzed to obtain the extent of counterion binding and the thermodynamic parameters of micellization. The values of delta H0, delta G0, and delta S0 obtained in 0.011 M Tris, pH 8.3, are -6 K X J X mol-1, -64 K X J X mol-1, and +193 J X mol-1 X K-1, and the average number of univalent ions bound per molecule in the micelles is 1.4. These values are within the range of those obtained for other univalent and polyvalent detergents. Analyzed by sedimentation and diffusion, the micelles are approximately spherical with an anhydrous mass of 50,000 daltons but with dimensions inconsistent with fully extended molecules. Correlation of the information obtained from the present physical studies with kinetic studies using long-chain fatty acyl-CoAs as enzyme substrates may be helpful for understanding the enzymology of these compounds, and some previously published kinetic studies of membrane-bound and soluble enzymes may bear reinterpretation.
Highlights
CoA were determined by surface tension, conductivitym, icelles
Are within the range of those obtained for otheruni- Until recently, the CMC of palmitoyl-CoA has been taken valentandpolyvalentdetergents.Analyzedby sedi- to be invariant at about 4 pM, an estimate obtained by using mentationanddiffusion,the micelles areapproxi- the cationic dye pinacyanol chloride (10) and subsequently mately spherical with an anhydrous mass of 50,000 verified by using the same probe or rhodamine 6G (11).In daltons but with dimensionsinconsistent with fully contrast, alternative methods such as analytical ultracentriextended molecules
Correlation of the information ofubg-ation, spin labeling with 6-doxylstearoyl-CoA,gel permeatained from the present physical studies with kinetic tion chromatography (61, and ultrafiltration (7) suggest that studies using long-chain fatty acyl-CoAs as enzyme the CMCof palmitoyl-CoA may be more than an order of esmliunsobhzlyseotmdgryaketsieonmsfemattiyhcaeysbsteebuaedcrhioeemrslepopifnfoutumlenrfepdomrsr,ebtuaranantniddoeen-rsb.sootmaunenddpinragenvdtihosueonslluzybyl-peubvcem-rnaeartpgiadantniiistocupnideasesriiithnnyibigtoehhnteewircmetsehetnraaestnnhugrepetrrhdee,vsCbuiMoultutsCstlfhsyreomdmiregatthethhrtemebsreeilnaaelrcadgct.etoeSaurnmtnedtacelhdclneofrionqrsduibiessyts
Summary
The critical micelle concentration (CMC’) is reached and association into micelles begins; abovethe CMC, the concentration of free molecules in equilibrium with micelles remains nearly constant and independentof total concentration providing mixedmicelles are not present (3) It would not be surprising if the pronounced change in physical state at theCMC werereflected as changes in kinetic parameters, and anomalous kinetic inhibition is frequently encountered when fatty acyl-CoAs are used as substrates (1, sign has been shown to introduce artifacts whenused to determine CMCs (12,13), andin the case of SDS-pinacyanol chloride the formation of a salt of the dye and detergent has been demonstrated to take place at detergent concentrations well below the CMC (12). We have employed this last method, together with the direct and relatively nonperturbing methods of surface tension and conductivity, to monitor the CMC of fatty acyl-
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