Editing Circular dichroism (section)

==Application to biological molecules==
[[File:Circular dichroism and structures calculation-protein in detergents.svg|thumb|400px|Upper panel: Circular dichroism spectroscopy in the far-ultraviolet wavelength region (far-UV CD) of MBP-cytochrome b<sub>6</sub> fusion protein in different detergent solutions. It shows that the protein in DM, as well as in Triton X-100 solution, recovered its structure. However the spectra obtained from SDS solution shows decreased ellipticity in the range between 200 and 210&nbsp;nm, which indicates incomplete secondary structure recovery. <br/>
Lower panel: The content of secondary structures predicted from the CD spectra using the CDSSTR algorithm. The protein in SDS solution shows increased content of unordered structures and decreased helices content.<ref>{{cite journal|author=Surma M.A.|author2=Szczepaniak A.|author3=Króliczewski J.|title=Comparative Studies on Detergent-Assisted Apocytochrome b6 Reconstitution into Liposomal Bilayers Monitored by Zetasizer Instruments|journal=PLOS ONE|volume=9|issue=11|year=2014|pages=e111341|issn=1932-6203|doi=10.1371/journal.pone.0111341|bibcode = 2014PLoSO...9k1341S|pmid=25423011|pmc=4244035|doi-access=free}}</ref>]]
Circular Dichroism (CD) spectroscopy is a powerful tool in biophysical and biochemical research, providing unique insights into the structural and stability characteristics of biomolecules. Because CD arises from the differential absorption of left- and right-circularly polarized light by [[optically active]] molecules, it is inherently sensitive to the chiral nature of biological macromolecules. This makes it particularly valuable for analyzing secondary structures, as seen in the characteristic CD spectral signatures of the [[Alpha helix|α-helices]] and [[Beta sheet|β-sheets]] of proteins and the double helices of [[nucleic acid]]s. While high-resolution techniques such as [[X-ray crystallography]], [[Nuclear magnetic resonance|NMR]], and [[Cryogenic electron microscopy|cryo-EM]] reveal atomic-level structural details, and interaction-based methods like [[Isothermal titration calorimetry|ITC]] and [[Surface plasmon resonance|SPR]] probe molecular interactions, CD spectroscopy offers a rapid, label-free means of detecting structural changes and stability shifts. Its ability to provide complementary data makes it an essential component of the modern biophysical toolbox, with applications spanning virtually every field of biomolecular research. CD spectroscopy is widely used in academia and the biopharmaceutical industry to study biomolecules, particularly proteins and peptides. CD spectra provide valuable insights into both [[Protein secondary structure|secondary]] and [[Protein tertiary structure|tertiary]] structure.

[[Protein secondary structure|Secondary structure]] information is derived from signals arising from peptide bonds that absorb in the far-UV range (spanning approximately 180&nbsp;nm to 260&nbsp;nm), enabling the identification and fractional assignments of structural elements such as the [[Alpha helix|alpha-helix]], [[Beta sheet|beta-sheet]], [[Beta turn|beta-turn]], and [[random coil]].<ref>{{cite journal |vauthors=Hall V, Nash A, Rodger A |year=2014 |title=SSNN, a method for neural network protein secondary structure fitting using circular dichroism data |url=http://wrap.warwick.ac.uk/75654/1/WRAP_9471544-ch-040116-140618_ssnn_a_method_for_neural_network_protein_secondary_structure_fitting_using_circular_dichroism_data_main.pdf |url-status=live |journal=Analytical Methods |volume=6 |issue=17 |pages=6721–26 |doi=10.1039/C3AY41831F |archive-url=https://ghostarchive.org/archive/20221009/http://wrap.warwick.ac.uk/75654/1/WRAP_9471544-ch-040116-140618_ssnn_a_method_for_neural_network_protein_secondary_structure_fitting_using_circular_dichroism_data_main.pdf |archive-date=2022-10-09}}</ref><ref>{{cite journal |vauthors=Hall V, Nash A, Hines E, Rodger A |year=2013 |title=Elucidating protein secondary structure with circular dichroism and a neural network |journal=Journal of Computational Chemistry |volume=34 |issue=32 |pages=2774–86 |doi=10.1002/jcc.23456 |pmid=24122928 |s2cid=19685126}}</ref><ref name="pmid17896349">{{cite journal |vauthors=Whitmore L, Wallace BA |year=2008 |title=Protein secondary structure analyses from circular dichroism spectroscopy: methods and reference databases |journal=Biopolymers |volume=89 |issue=5 |pages=392–400 |doi=10.1002/bip.20853 |pmid=17896349 |doi-access=}}</ref><ref name="pmid17406547">{{cite journal |author=Greenfield NJ |year=2006 |title=Using circular dichroism spectra to estimate protein secondary structure |journal=Nature Protocols |volume=1 |issue=6 |pages=2876–90 |doi=10.1038/nprot.2006.202 |pmc=2728378 |pmid=17406547}}</ref> These structural assignments place important constraints on the possible secondary conformations that the protein can be in. CD cannot, in general, say where the alpha helices that are detected are located within the molecule or even completely predict how many there are. Despite this, CD is a valuable tool, especially for showing changes in conformation. It can, for instance, be used to study how the secondary structure of a molecule changes as a function of temperature (in a thermal denaturation temperature ramp experiment) or of the concentration of denaturing agents (in a chemical denaturation experiment), e.g. [[guanidinium chloride]] or [[urea]]. In this way it can reveal important thermodynamic information about the molecule (such as the [[enthalpy]] and [[Gibbs free energy]] of denaturation) that cannot otherwise be easily obtained. Anyone attempting to study a protein will find CD a valuable tool for verifying that the protein is in its native conformation (i.e., as a quality control technique) before undertaking extensive and/or expensive experiments with it. Moreover, CD spectroscopy has been used in [[Bioinorganic chemistry|bioinorganic]] interface studies; specifically, it has been used to analyze the differences in secondary structure of an engineered protein before and after titration with a reagent.<ref name="pubs.acs.org">Bioinorganic Interface: Mechanistic Studies of Protein-Directed Nanomaterial Synthesis. (2016, May 5). Retrieved March 1, 2019, from https://pubs.acs.org/doi/pdf/10.1021/acs.jpcc.6b02569</ref>

Insight into [[Protein tertiary structure|tertiary structure]] is obtained from spectral contributions of [[aromatic amino acid]]s, which absorb in the near-UV range (spanning approximately 250&nbsp;nm to 350&nbsp;nm). The signals obtained in this region are due to the absorption, dipole orientation, and nature of the surrounding environment of the phenylalanine, tyrosine, cysteine (or S-S [[disulfide bridges]]) and tryptophan [[amino acid]]s. Unlike in far-UV CD, the near-UV CD spectrum cannot be assigned to any particular 3D structure. Rather, the near-UV range can be considered a fingerprint region because its spectral profile is exquisitely dependent on the composition of aromatic residues and the conformation and environment of their side chains.

Visible CD spectra (spanning approximately 350&nbsp;nm to 700&nbsp;nm) provide structural information on the nature of the prosthetic groups in proteins, e.g., the heme groups in [[hemoglobin]] and [[cytochrome c]]. Visible CD spectroscopy is a very powerful technique to study metal–protein interactions and can resolve individual d–d electronic transitions as separate bands. CD spectra in the visible light region are generally only produced when a metal ion is in a chiral environment, thus, free metal ions in solution are not detected. This has the advantage of only observing the protein-bound metal, so pH dependence and stoichiometries are readily obtained. Optical activity in transition metal ion complexes have been attributed to configurational, conformational, and vicinal effects. Klewpatinond and Viles (2007) have produced a set of empirical rules for predicting the appearance of visible CD spectra for Cu(II) and Ni(II) square-planar complexes involving histidine and main-chain coordination.<ref>{{Cite journal |last1=Rodikova |first1=Ekaterina A. |last2=Kovalevskiy |first2=Oleg V. |last3=Mayorov |first3=Sergey G. |last4=Budarina |first4=Zhanna I. |last5=Marchenkov |first5=Victor V. |last6=Melnik |first6=Bogdan S. |last7=Leech |first7=Andrew P. |last8=Nikitin |first8=Dmitri V. |last9=Shlyapnikov |first9=Michael G. |last10=Solonin |first10=Alexander S. |date=2007-03-20 |title=Two HlyIIR dimers bind to a long perfect inverted repeat in the operator of the hemolysin II gene from Bacillus cereus |url=https://pubmed.ncbi.nlm.nih.gov/17346714/ |journal=FEBS Letters |volume=581 |issue=6 |pages=1190–1196 |doi=10.1016/j.febslet.2007.02.035 |issn=0014-5793 |pmid=17346714|bibcode=2007FEBSL.581.1190R }}</ref>

Beyond proteins and peptides, UV-CD spectroscopy is a powerful tool for characterizing the secondary structure of nucleic acids, providing insight into DNA helices and RNA structural motifs such as G-quadruplexes. Different DNA conformations—A-DNA, B-DNA, and Z-DNA—exhibit distinct CD spectral signatures due to variations in base stacking and helical geometry.<ref>{{Cite journal |last1=Baker |first1=Erin Shammel |last2=Bowers |first2=Michael T. |date=2007-07-01 |title=B-DNA Helix Stability in a Solvent-Free Environment |url=https://link.springer.com/article/10.1016/j.jasms.2007.03.001 |journal=Journal of the American Society for Mass Spectrometry |language=en |volume=18 |issue=7 |pages=1188–1195 |doi=10.1016/j.jasms.2007.03.001 |pmid=17434745 |bibcode=2007JASMS..18.1188B |issn=1879-1123}}</ref><ref>{{Cite journal |last1=Kypr |first1=Jaroslav |last2=Kejnovská |first2=Iva |last3=Renčiuk |first3=Daniel |last4=Vorlíčková |first4=Michaela |date=2009-04-01 |title=Circular dichroism and conformational polymorphism of DNA |url=https://academic.oup.com/nar/article/37/6/1713/1030465 |journal=Nucleic Acids Research |volume=37 |issue=6 |pages=1713–1725 |doi=10.1093/nar/gkp026 |issn=0305-1048}}</ref> Similarly, RNA structures, including stem-loops, pseudoknots, and G-quadruplexes, produce unique CD spectra that reflect their specific folding patterns and base interactions. G-quadruplexes, in particular, show characteristic positive and negative bands in the CD spectrum depending on their topology (parallel, antiparallel, or hybrid).<ref>{{Cite journal |last1=del Villar-Guerra |first1=Rafael |last2=Trent |first2=John O. |last3=Chaires |first3=Jonathan B. |date=2018 |title=G-Quadruplex Secondary Structure Obtained from Circular Dichroism Spectroscopy |journal=Angewandte Chemie International Edition |language=en |volume=57 |issue=24 |pages=7171–7175 |doi=10.1002/anie.201709184 |issn=1521-3773 |pmc=5920796 |pmid=29076232}}</ref> These spectral features make UV-CD an essential technique for studying nucleic acid folding, stability, and interactions with ligands.

CD gives less specific structural information than [[X-ray crystallography]] and [[protein NMR]] spectroscopy, for example, which both give atomic resolution data. However, CD spectroscopy is a quick method that does not require large amounts of proteins or extensive data processing. Thus CD can be used to survey many [[solvent]] conditions, varying [[temperature]], [[pH]], [[salinity]], and the presence of various cofactors.

CD spectroscopy is usually used to study proteins in solution, and thus it complements methods that study the solid state. This is also a limitation, in that many proteins are embedded in [[biological membrane|membranes]] in their native state, and solutions containing membrane structures are often strongly scattering. CD can also be measured in thin films and powders. For instance, CD spectroscopy has been conducted on solid state semiconducting materials such as TiO<sub>2</sub> to obtain large signals in the UV wavelength range, where the electronic transitions for biomolecules often occur.<ref>Sarkar, Sumant, Ryan Behunin, and John G. Gibbs. "Shape-Dependent, Chiro-Optical Response of UV-Active, Nanohelix Metamaterials." Nano letters (2019). https://pubs.acs.org/doi/10.1021/acs.nanolett.9b03274</ref>