Identification of Small-Molecule Noncovalent Binders Utilizing SAMDI Technology

Area of Science:

• Analytical Chemistry
• Biochemistry
• Drug Discovery

Background:

• Mass spectrometry (MS) is increasingly vital in drug discovery for analyzing complex mixtures.
• High-throughput screening (HTS) requires robust assays, especially for targets lacking alternative methods.
• Label-free MS offers high accuracy and resolving power for analyte identification.

Purpose of the Study:

• To present a novel screening format using Self-Assembled Monolayers and matrix-assisted laser Desorption Ionization (SAMDI) technology.
• To demonstrate SAMDI's utility for label-free, high-throughput screening of small-molecule interactions.
• To validate SAMDI's performance in affinity capture, noncovalent interaction identification, and binding affinity assessment.

Main Methods:

• Utilized Self-Assembled Monolayers (SAM) for affinity capture of target proteins.
• Employed matrix-assisted laser Desorption Ionization (MALDI) time-of-flight mass spectrometry for ionization and detection.
• Implemented a pooled library format for enhanced screening throughput.
• Minimized variability using uniformly saturated protein surfaces to avoid crystallization hotspots.

Main Results:

• SAMDI technology enabled label-free identification of noncovalent compound interactions.
• Reduced interference from salts, detergents, and matrix through high-resolution MS.
• Achieved improved screening throughput irrespective of enzyme activity using pooled libraries.
• Demonstrated correlation between binding affinity rank ordering and biochemical assay potency data.

Conclusions:

• SAMDI provides a robust and high-throughput platform for MS-based drug discovery screening.
• The technology overcomes common challenges in MS screening, such as shot-to-shot variability and interference.
• SAMDI effectively identifies and ranks small-molecule binders, supporting early-stage drug development.