Protein Analysis/HOS
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Analysis of proteins and high order structures in drug development

The complex structure of proteins, their interactions and their various post-translational modifications (PTMs) make studying their actions and relationships a significant challenge. Broad protein analysis and characterisation is required during drug development, to achieve a comprehensive understanding of a number of characteristics, including the protein’s structure, physiochemical properties, biological activity and impurities.

Confirmation of protein identity through structural characterisation facilitates the understanding of the molecule’s molecular structure, from the primary amino acid sequence to the high order structure. Researchers want to be able to isolate structural effects of a protein during drug binding or interaction with substrates/inhibitors.

Disulfide bond (DSB) analysis and hydrogen deuterium exchange (HDX) have emerged as technologies that can be employed to gain insights into protein structure. These insights are critical for determining structural similarity for biosimilars, or for monitoring protein stability during drug development. The tertiary structure of therapeutic proteins is key to their activity and stability. 

Research and development labs require technology capable of automated DSB analysis in biopharmaceuticals, based on a single digest of the unreduced protein and without prior knowledge of enzyme specificity or native DSBs. Due to the complexity of these proteins and the fact that they will contain multiple disulfide bonds, analysis is a challenge, often requiring several LC-MS runs with the tryptic digests from reduced and non-reduced protein and a manual comparison of these two analyses.

Fourier Transform Infrared (FT-IR) spectroscopy can be used to analyse water-soluble and membrane-bound proteins, the latter of which are the main targets in drug research and development. Fast data acquisition and high sample throughput are some of the benefits of this technique. Infrared protein analysis is also relatively inexpensive and is a powerful technique used for formulation optimisation, stability studies during drug development and QC of protein drug products. 

Another technique used for the study of membrane proteins is surface plasmon resonance (SPR) – an optical detection method – which accurately and rapidly measures the binding processes in the analysis of biomolecular interactions. Some technologies have been developed to combine SPR with mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy, enabling the examination of bound molecules.