Membrane proteins are some of the most valuable targets in drug discovery, accounting for over half of all therapeutic targets. However, their inherent structural diversity and the complexity of their preparation often make them difficult to work with in screening campaigns.
Membrane proteins are at the start of many signaling pathways and are often present at the cell surface. These factors make them easier to access with biologic and small molecule drugs, explaining their over-representation as drug targets.
Unlocking the therapeutic potential of membrane proteins is challenging due to several inherent features. They are significantly less abundant than most soluble proteins and are often only expressed in a particular cell type or tissue. Being native to a phospholipid bilayer, extraction with detergents is typically required to isolate the protein, which can result in unfolding or aggregation.
Affinity selection-mass spectrometry (AS-MS) offers a powerful, tag-free approach to identify membrane proteins that does not require immobilization, minimizing conformational limitations. In this interview, Dr. Renaud Prudent, scientific director at Edelris, discusses how AS-MS can be used as a high-throughput screening technology for membrane protein hit identification.
Blake Forman (BF):
Senior Science Writer
Technology Networks
Blake pens and edits breaking news, articles and features on a broad range of scientific topics with a focus on drug discovery and biopharma. He earned an honors degree in chemistry from the University of Surrey. Blake also holds an MSc in chemistry from the University of Southampton. His research project focused on the synthesis of novel fluorescent dyes often used as chemical/bio-sensors and as photosensitizers in photodynamic therapy.
Membrane proteins are notoriously difficult to work with in screening campaigns. From your perspective, what makes AS-MS particularly well-suited for tackling these challenges?
Scientific Director
Edelris
Dr. Renaud Prudent is the scientific director at Edelris, where he has implemented AS-MS screening technologies since joining in 2018. Prudent holds a PhD in chemical biology from Université Grenoble Alpes, where he specialized in screening allosteric kinase inhibitors.
We know that membrane proteins can oscillate between different conformations. Many AS-MS methods utilize the sample in solution, so we don’t restrict this conformational landscape. We can also use untagged proteins, which may bypass the issues that come with using tags. AS-MS encompasses a range of techniques that can be adapted based on the profile of the protein of interest.
BF:
Senior Science Writer
Technology Networks
Blake pens and edits breaking news, articles and features on a broad range of scientific topics with a focus on drug discovery and biopharma. He earned an honors degree in chemistry from the University of Surrey. Blake also holds an MSc in chemistry from the University of Southampton. His research project focused on the synthesis of novel fluorescent dyes often used as chemical/bio-sensors and as photosensitizers in photodynamic therapy.
Traditional screening methods often focus on specific functional outcomes. How does the activity-agnostic nature of AS-MS change the way researchers can approach target discovery and characterization?
RP:
Scientific Director
Edelris
Dr. Renaud Prudent is the scientific director at Edelris, where he has implemented AS-MS screening technologies since joining in 2018. Prudent holds a PhD in chemical biology from Université Grenoble Alpes, where he specialized in screening allosteric kinase inhibitors.
The AS-MS detects binders and is not dependent on functional outcome. This enables us to detect and characterize agonists, antagonists and allosteric binders. AS-MS can also be used to detect silent binders – a compound that binds to the protein with no functional effect on that target's activity – which may be of interest in protein target degradation or protein-protein interactions. This is why, to some extent, AS-MS varies from traditional screening techniques, as we can detect things that other techniques cannot. This feature makes it especially useful for discovering chemical modulators with diverse mechanisms of action.
In terms of sensitivity, the increase in power of mass spectrometers makes it possible to detect binder from the nanomolar to the two- to three-digit micromolar range.
Senior Science Writer
Technology Networks
Blake pens and edits breaking news, articles and features on a broad range of scientific topics with a focus on drug discovery and biopharma. He earned an honors degree in chemistry from the University of Surrey. Blake also holds an MSc in chemistry from the University of Southampton. His research project focused on the synthesis of novel fluorescent dyes often used as chemical/bio-sensors and as photosensitizers in photodynamic therapy.
How adaptable is AS-MS across diverse targets?
RP:
Scientific Director
Edelris
Dr. Renaud Prudent is the scientific director at Edelris, where he has implemented AS-MS screening technologies since joining in 2018. Prudent holds a PhD in chemical biology from Université Grenoble Alpes, where he specialized in screening allosteric kinase inhibitors.
In AS-MS, the detection is made by a mass spectrometer, but the affinity selection component comes in a range of guises. For example, size exclusion assays can be conducted through liquid chromatography. You also have filter-binding assays and the pull-down assays (where you need a tagged target). Depending on the target's size and stability, and whether you want to screen a complex or focus on a specific binding site, you can tailor the affinity selection and evaluate this experimentally before screening.
BF:
Senior Science Writer
Technology Networks
Blake pens and edits breaking news, articles and features on a broad range of scientific topics with a focus on drug discovery and biopharma. He earned an honors degree in chemistry from the University of Surrey. Blake also holds an MSc in chemistry from the University of Southampton. His research project focused on the synthesis of novel fluorescent dyes often used as chemical/bio-sensors and as photosensitizers in photodynamic therapy.
What do you see as some of the biggest barriers to broader adoption of AS-MS in drug discovery pipelines, and how might the field overcome them?
RP:
Scientific Director
Edelris
Dr. Renaud Prudent is the scientific director at Edelris, where he has implemented AS-MS screening technologies since joining in 2018. Prudent holds a PhD in chemical biology from Université Grenoble Alpes, where he specialized in screening allosteric kinase inhibitors.
If you compare AS-MS with standard screening, the main barrier is data processing. We have many mass spectra to analyze to pinpoint which component binds to the target. If you compare this with a fluorescent assay, where you read the plate and can monitor the result, there's a lot of processing required to interpret what's going on. We’ve spent quite a lot of time trying to streamline the data analysis and overcome this.
Currently, the classical algorithm works quite well, but AI will enable us to focus on the most trusted binder by comparing it with internal data sets.
Another area we are looking to improve is component management. We need to have a large library size (more than 2 million compounds), and you must consider mass redundancy to determine what the optimal pool is. Additionally, there is an initial cost associated with setting up the dedicated platform, which is typically higher in comparison to fluorescence techniques. However, this cost is often offset by the high-throughput screening capabilities of AS-MS. For example, in 1 well, we can screen 500 compounds, and we are working on improving this further.
Senior Science Writer
Technology Networks
Blake pens and edits breaking news, articles and features on a broad range of scientific topics with a focus on drug discovery and biopharma. He earned an honors degree in chemistry from the University of Surrey. Blake also holds an MSc in chemistry from the University of Southampton. His research project focused on the synthesis of novel fluorescent dyes often used as chemical/bio-sensors and as photosensitizers in photodynamic therapy.
Looking ahead, what future applications or technological advances do you anticipate will further expand the role of AS-MS in studying membrane proteins and beyond?
RP:
Scientific Director
Edelris
Dr. Renaud Prudent is the scientific director at Edelris, where he has implemented AS-MS screening technologies since joining in 2018. Prudent holds a PhD in chemical biology from Université Grenoble Alpes, where he specialized in screening allosteric kinase inhibitors.
The current trend is targeted protein degradation for which AS-MS fits well since we can detect binders that are used as warheads for the proteins of interest. We are also developing assays to identify molecular glues, which are compounds that allow two proteins to stick together. Another trend is the integration of not just isolated proteins, but more complex biological systems that provide access to more physiologically relevant information.
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