Secondary structure assessment of formulated bevacizumab in the presence of SDS by deep ultraviolet resonance Raman (DUVRR) spectroscopy

摘要

A deep-ultraviolet resonance Raman (DUVRR) spectroscopic method has been used to study the secondary structural changes of a therapeutic monoclonal antibody (mAb), bevacizumab (Avastin?) under a chemical stress: the presence of sodium dodecyl sulfate (SDS). The results demonstrate that DUVRR spectroscopy can assay the higher order structure of the formulated protein in a sensitive and selective manner. The SDS-induced partially unfolding of the mAb was probed by DUVRR spectroscopy where the amide I, II and III spectral features showed conformational changes between beta-sheet, alpha-helix and random coil forms. A chemometric model was also built to analyze the spectral changes occurring with protein-SDS interactions. The analysis showed there are different stages of mAb-SDS interaction as the SDS concentration increases. In addition, a two-dimensional (2D) correlation analysis was applied to the DUVRR spectra to visualize the secondary structure changes of bevacizumab under stresses. As an addition to the chemometric model, the 2D correlation mapping method suggested different transitions between secondary structure motifs were occurring at different SDS concentrations. Overall, chemometric and 2D analysis provided complimentary information, and show the potential of coupling DUVRR with advanced statistical methods in revealing complex structural information in formulated protein pharmaceuticals.