Circular Dichroism Spectra of α-Chymotrypsin–SDS Solutions Depend on the Procedure of Their Preparation

Abstract

We recorded the far- and near-UV circular dichroism (CD) spectra of solutions of α-chymotrypsin and sodium dodecyl sulfate (SDS) with the final surfactant concentration significantly above the critical micellization concentration. Solutions were prepared using three different procedures. The reference procedure was to mix the chymotrypsin solution with the SDS solution once, immediately achieving the final SDS concentration. In alternative procedures, the protein solutions initially contained some SDS and were mixed with pure SDS solutions at a concentration to provide the same final surfactant as the reference mixing. We demonstrate that the supplementation to the selected final concentration of SDS of the pure chymotrypsin solution leads to different CD spectra than the supplementation to this final concentration of SDS in the chymotrypsin solution containing a small concentration of a few millimolar SDS. These differences disappear when the initial concentration of SDS in the protein solution, which we then supplement to the indicated final concentration, is higher. This suggests the irreversibility of the processes caused by the addition of SDS to chymotrypsin and the influence of the initial amount of this surfactant on the processes occurring with its further addition to the solution. For quantitative analysis of far-UV CD spectra in terms of populations of protein secondary structure elements, we used four well-established software packages. All programs consistently indicate that the observed differences in the far-UV CD spectra can be explained by the differences in the increase in the population of helical forms in chymotrypsin under the influence of SDS.

Kinetics of Structural Transitions Induced by Sodium Dodecyl Sulfate in α-Chymotrypsin

“…The results shown in Figure indicate that the final protein state obtained after mixing with SDS depends not only on the final concentration of the surfactant but also on the mixing procedure. These different final states seem relatively stable, as confirmed by Figure and our previous investigations …”

“…These different final states seem relatively stable, as confirmed by Figure 11 and our previous investigations. 55 …”

“…These different final states seem relatively stable, as confirmed by Figure 11 and our previous investigations. 55 Figure 11 shows the stationary CD spectra obtained after mixing the 40 μM CHA + 4 mM SDS solution with the solution of 76 mM SDS. These spectra are stable for at least 4 h after the mixing.…”

Interaction Mechanism between α-Lactalbumin and Caffeic Acid: A Multispectroscopic and Molecular Docking Study

“…Far-UV CD is a very good spectroscopic technique used in biology and chemistry to determine the changes in the protein secondary structure . Spectra in the far UV range of ∼200–250 nm provide detailed information on protein polypeptide backbone conformations.…”

“…Far-UV CD is a very good spectroscopic technique used in biology and chemistry to determine the changes in the protein secondary structure. 42 Spectra in the far UV range of ∼200–250 nm provide detailed information on protein polypeptide backbone conformations. Intermolecular forces that are responsible for holding secondary and tertiary protein structures are observed during ligand interactions with protein molecules, which further aid in protein conformational changes.…”