Gain insight into secondary structure
Typical beta-sheet structure retained in native and stressed mAb.
Disulphide bond environment unchanged.
Spectra normalized for protein concentration by simultaneous absorbance measurements, 0.5 mm pathlength, Chirascan™ V100
Spectra normalized for protein concentration by simultaneous absorbance measurements, 10 mm pathlength, Chirascan™ V100
Reveal tertiary structure
Environment of aromatic amino acid side-chains appears similar.
Note: A subsequent HOS comparison study using a Chirascan™ Q100 system confirmed that minor differences in tertiary structure were statistically significant (data not shown).
Conclusion
Review of spectra suggests no difference between native and stressed mAb.
Effect of transient exposure to nanoparticles
(Data courtesy of a leading research university, Germany)
Spectra normalized for protein concentration by simultaneous absorbance measurements, 0.5 mm pathlength, Chirascan™ V100
Determine structural effects
Significant change in far UV CD spectrum of an enzyme indicated perturbation of secondary structure by nanoparticle exposure
Continuous temperature ramp 20°- 90°C, 71 spectra in 71 min, 1°C/min, 0.8 sec per point, 1 nm bandwidth, 1 nm step, Chirascan™ V100
Determine effect on protein stability
Thermal denaturation curves showed altered folding profile after nanoparticle exposure. Global fit of multiwavelength data confirmed a 2.9°C change in melting temperature.
Publications
Since the introduction of the first instrument in 2005, Chirascan CD spectrometers have contributed to thousands of peer-reviewed publications. Below are featured a small selection of the most recent publications showing the breadth of applications.
Austerberry et al. 2017. “The Effect of Charge Mutations on the Stability and Aggregation of a Human Single Chain Fv Fragment.” European Journal of Pharmaceutics and Biopharmaceutics 115 (June): 18–30.
Bui and Nguyen. 2016. “Insights Into the Interaction of CD4 with Anti-CD4 Antibodies.” Immunobiology 222 (2): 148–54.
Matukumalli, Tangirala, and Rao. 2017. “Clusterin: Full-Length Protein and One of Its Chains Show Opposing Effects on Cellular Lipid Accumulation.” Scientific Reports 7 (January). Nature Publishing Group: 41235.
Miao et al. 2017. “Physicochemical and Biological Characterization of the Proposed Biosimilar Tocilizumab.” BioMed Research International 2017 (March). Hindawi: 1–13.
Moreno et al. 2016. “Study of Stability and Biophysical Characterization of Ranibizumab and Aflibercept.” European Journal of Pharmaceutics and Biopharmaceutics 108: 156–67.
Narang, Singh, and Mukhopadhyay. 2017. “Stepwise Unfolding of Human β2-Microglobulin Into a Disordered Amyloidogenic Precursor at Low pH.” European Biophysics Journal 46 (1). Springer Berlin Heidelberg: 65–76.
Ropiak et al. 2017. “Identification of Structural Features of Condensed Tannins That Affect Protein Aggregation.” Edited by Jamshidkhan Chamani. PLOS ONE 12 (1): e0170768.
Townsend et al. 2017. “Heparin and Methionine Oxidation Promote the Formation of Apolipoprotein A-I Amyloid Comprising α-Helical and β-Sheet Structures.” Biochemistry 56 (11). American Chemical Society: 1632–44.
Vorob’ev et al. 2017. “Physicochemical Properties, Toxicity, and Specific Activity of a Follitropin Alpha Biosimilar.” Pharmaceutical Chemistry Journal 50 (11). Springer US: 753–60.
Zhao et al. 2016. “Structural Insights on PHA Binding Protein PhaP from Aeromonas Hydrophila.” Scientific Reports 6 (1): 39424.