Abstract

Proteins are complex macromolecules that require comprehensive analysis. High resolution techniques are used to determine structure of proteins, yet further understanding of protein dynamics and behavior is needed for understanding protein stability. Deamidation is a phenomenon that occurs spontaneously in proteins, while in solution when subject to heat stress, low or high pH. The mechanism of deamidation is known, involving an asparagine residue undergoing a succinimide intermediate resulting in the generation of an aspartate residue. Implications of charge in the protein of interest due to a deamidation can lead to events such as aggregation, loss of affinity towards its target, and truncation. We have developed an innovative Best-in-Class label free high throughput platform technology specifically designed for comparative characterization of an array of proteins in solution. This platform technology takes advantage of: (1) Quantum Cascade Laser (QCL) microscope with enhanced signal-to-noise ratio (SNR) within the MID IR spectral region 1780 - 1000 cm−1; (2) innovative cells, (3) Hyperspectral Images (HSI) that allow for the visual inspection of the sample to determine the size and identity of any aggregates in the 4.3 µm - 2 mm size range and (4) QCL IR spectral analysis providing amino acid level resolution that is used to define the molecular behavior of proteins in solution. Our Correlation Dynamics software with 2D IR and Co-distribution correlation algorithms are used to map glutamine and asparagine deamidation sites or assess domain stability have a direct impact on drug design and development. Furthermore, ProteinMentor is well suited to be implemented orthogonally with other well-established analytical techniques. Two case studies will be presented as proof-of-concept and validation: (1) NIST mAb standard, subject to Hurricane Maria and (2) NCI mAb fragments.

Full Text
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