Circular Dichroism Method for Heat Capacity Determination of Proteins

Abstract

Circular dichroism spectroscopy was used to measure the thermal unfolding of bovine pancreatic ribonuclease A (RNase A) with various concentrations of guanidine hydrochloride (GuHCl). A red shift in transition midpoint temperatures, T m, occurred with increasing concentration of the strong protein denaturant. van Hoff enthalpy changes, ΔH°, were calculated and plotted as a function of T m to determine the heat capacity change, ΔC p, for denaturation. A value of 4.02 ± 0.02 kJ mol–1 K–1 was calculated from d( ΔH)/d(ΔT m). Reported values for ΔC p range from 4.2 to 9.6 kJ mol–1 K–1. The shift in T m for RNase A with increasing concentration of GuHCl suggests that the protein is undergoing substantial changes in its secondary structure.

Circular Dichroism of G-Quadruplex: a Laboratory Experiment for the Study of Topology and Ligand Binding

“…The application of CD spectroscopy in the classroom has been previously described, particularly to the study of proteins. − In this laboratory experiment we addressed the study of noncanonical DNA structures, namely, G4s.…”

“…The analysis of CD spectra is currently part of the practice in structural analysis and structural biochemistry of undergraduate and graduate courses for biochemistry, biomedicine, and medicinal chemistry degrees, and has been a subject of publication in a variety of education-related articles. − …”

Circular Dichroism Spectroscopy in the Undergraduate Curriculum

“…Numerous protocols have been published on the use of CD spectroscopy in the classroom, including several from this Journal − . Some aim to introduce CD to students as an unusual method for measuring common chemical parameters (e.g., p K a ) , but most use the unique features of CD as a spectral polarimetry technique that enables confirmation of the spectral fingerprint of protein structure , , measurement of the stability or heat capacity of a protein, determination of absolute configuration , , or monitoring the synthetic conversion of one chiral organic compound into another .…”

“…Numerous protocols have been published on the use of CD spectroscopy in the classroom, including several from this Journal − . Some aim to introduce CD to students as an unusual method for measuring common chemical parameters (e.g., p K a ) , but most use the unique features of CD as a spectral polarimetry technique that enables confirmation of the spectral fingerprint of protein structure , , measurement of the stability or heat capacity of a protein, determination of absolute configuration , , or monitoring the synthetic conversion of one chiral organic compound into another . These examples highlight the simple yet powerful way in which CD spectroscopy can be used to demonstrate basic concepts, such as molecular asymmetry, as well as advanced concepts, such as the thermodynamic parameters that govern protein folding.…”

Electrochemical Deposition in RF-Irradiated Solutions - A Manifestation of Induced Long Range Order?

“…The CD spectroscopy is widely used to investigate proteins properties such as folded/open structure, secondary and tertiary structures (29). Commonly the temperature of the measured solution can be varied thus enabling to detect the kinetic of structure changes such as thermal unfolding of proteins (30). The CD is also used to clarify conformation of DNA in liquid/solid interface (31), determined self-assembles of polymeric nanoparticles (32) and investigate molecular chirality in general (33).…”