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Higher Order Structures: Gain Insights to Protein Structure

Disulfide bond (DSB) analysis and hydrogen deuterium exchange (HDX) have emerged as technologies that can be employed to gain insights to protein structure. These insights are critical for determining structural similarity for biosimilars, or for monitoring protein stability during innovator development.

Disulfide bond analysis with DisulfideDetect

The tertiary structure of therapeutic proteins is key to their activity and stability. 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.

The DisulfideDetect workflow allows the fully automated analysis of disulfide bonds in biopharmaceuticals, based on a single digest of the unreduced protein and without prior knowledge of enzyme specificity or native disulfide bonds. The workflow also provides a semi-quantitative scrambling analysis as well.

DisulfideDetect software sets a new standard in ease-of-use for detecting and evaluating disulfide bonds within therapeutic proteins, such as monoclonal antibodies. It provides an easy to understand, visually-based, workspace for disulfide bond analysis for both native and non-native bonds as well as inter- and intra-chain disulfide bonds within the protein/antibody.

DisulfideDetect software uses established methods from existing LC-MALDI instruments like the UltrafleXtremeTM, so if your lab is utilizing LC-MALDI you're all set! Just add the DisulfideDetect Module to begin analyzing disulfide bonds in your biotherapeutics. No additional hardware is necessary.

 

 

Hydrogen Deuterium Exchange

The biopharmaceutical industry is increasingly required to provide data illustrating any structural changes and interactions a therapeutic drug might undergo, and the effects these could have on the stability and efficacy of the drug. HDX can be used to study structural hotspots, perform epitope mapping studies and to monitor the effects of stressors on protein structure.

The Bruker HDX Solution combines OTOF mass spectrometry with the most advanced HDX automation platform available today, the Leap H/D-X PAL liquid handling robot. This robust and proven system is controlled by Chronos software, which optimizes sample scheduling for increased efficiency and productivity. HDExaminer software (Sierra Analytics) extracts the HDX information from the MS data, converting data into knowledge of changes in protein conformation.   

Bruker’s OTOF mass spectrometers provide sub-ppm mass accuracy, high sensitivity and True Isotopic Patterns (TIP TM) which are all essential for generating and quantifying hydrogen/deuterium ratios with the highest confidence.