Synchrotron radiation circular dichroism spectroscopy

The physical principles of SRCD are essentially identical to those of CD, in that the technique measures the difference in absorption (ΔA) of left (AL) and right (AR) circularly polarized light (ΔA=AL-AR) by a sample in solution.

The major advantages of SRCD over CD arise from the ability to measure data over an extended wavelength range into the vacuum ultra violet (VUV) end of the spectrum.

As these measurements are utilizing a light source with a higher photon flux (quantity of light stricking a given surface area) than a bench-top CD machine it means data are more accurate at these extended wavelengths because there is a larger signal over the background noise (the signal-to-noise ratio) and, generally, less sample is needed when recording the spectra and there is more information content available in the data.

[2] Synchrotron radiation (SR) had been proposed for use as the light source at a meeting in Brookhaven National Laboratory on Long Island in 1972,[2][3] however, it took a few years more before this came to fruition.

The use of only one mirror minimizes the loss of photon flux which is most important in the VUV region where reflectivity is poor relative to the visible wavelength range.

One of the primary factors limiting the lower wavelength cut off is the sample usually being in solution as a large water absorption band exists centred ~167 nm.

[11][12] Removing the solvating water completely, creating a film as a result, means that data can be recorded to significantly lower wavelengths, down to around ~130 nm.

By contrast, typically, the photon flux for an SRCD beamline in this region is at least three orders of magnitude higher than a cCD machine, retaining that level down to ~150 nm.

[15] While the first reports of its use dated to 1980, it was a further two decades before the technique of SRCD took off largely due to the work of Bonnie Wallace at Birkbeck College, University of London.

From around 2000, her aims in the field focused on both enhancing the collection of quality data through technical improvements, and on demonstrating "proof-of-principle" application studies, illustrating the novel information that SRCD offers.

[29][30][31] Additionally, her lab produced sample cells with reduced pathlengths, and using material, (CaF2), transparent to VUV radiation which significantly enhanced the collection of data into the SRCD lower wavelength regions.

SRCD investigations were conducted on the wild-type protein and two variants, the P23T mutant found in the disease, and a related modification, P23S (proline to serine, a chemically similar amino acid to threonine), to establish the nature of the cause of cataract formation.

[39] Because of a high degree of flexibility, it had proven difficult to determine the structure of the extramembranous C-terminal domain of bacterial voltage-gated sodium channels.

Using a series of synthesised channels where this C-terminal domain had been truncated, in some cases by a single amino acid difference between the constructs, the Wallace lab used SRCD to successfully identify the structure of this region.

IDPs (and IDRs if capable of being isolated from the rest of the protein) have a distinct SRCD spectral appearance in solution which means that changes in their spectra that arise through interactions offer an ideal opportunity to gain insight into what is happening both structurally and functionally.

In addition, SRCD studies have demonstrated that when the solvating water is removed from these proteins, generating a film, there is a gain in structure and more CD transition bands can be measured into the lower VUV wavelength region because the water absorption band is not present[43] Myelin is the insulating sheath that is formed in the central (CNS) and peripheral nervous systems (PNS) to surround nerve cell axons thereby increasing and maintaining the electrical impulse, the action potential, sent along them.

MBP and P0ct were employed in a study[44] which used SRCD data as a key factor to establish if there was any significance to the predictions of their IDP and IDR protein structures generated by Alphafold2, an artificial intelligence program developed by DeepMind.

[44] One major feature found in protein structures is the addition of sugars (glycosylation) to specific amino acid residues by post translational modification.