Our company has established a comprehensive bioactive natural products target identification technology platform, with activity-based protein profiling (ABPP) and click chemistry–ABPP (CC-ABPP) as core technologies. CC-ABPP integrates the selectivity of activity-based probes with the efficiency and bioorthogonality of click chemistry, enabling robust, sensitive, and unbiased identification of functional protein targets in complex biological systems.
Introduction to CC-ABPP
While traditional ABPP is powerful, it often requires attaching large reporter groups (like biotin or fluorophores) directly to the bioactive molecule. These bulky groups can hinder cellular uptake or mask the molecule's ability to bind to its target protein. CC-ABPP, or "two-step" ABPP, revolutionizes this process. Instead of a large tag, it incorporates a tiny, bioorthogonal "click" handle—typically an alkyne or an azide. This minimal modification preserves the natural product's size, lipophilicity, and biological activity. After the probe binds to its target in a living cell or lysate, the reporter group is "clicked" on via a Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) for detection and enrichment.
Fig. 1. Principle of CC-ABPP [1].
Our CC-ABPP Technology Platform
Our CC-ABPP technology platform is designed to handle the inherent complexity of natural products, which often feature intricate stereochemistry and multiple reactive centers. We provide a comprehensive suite of services that transform a bioactive "hit" into a validated "target," empowering researchers to understand the MoA (Mechanism of Action) at a granular level.
Our Probe Design Capabilities
The success of CC-ABPP hinges on the design of the probe. Our expert medicinal chemists specialize in:
- SAR-guided Probe Design: We carefully select labeling positions based on SAR data to minimize disruption of target binding.
- Minimalist Tagging: Identifying the optimal position to insert an alkyne or azide handle without disrupting the pharmacophore.
- Photo-affinity Labeling (PAL): For non-covalent natural products, we design bifunctional probes containing both a click handle and a photo-reactive group (e.g., diazirine or benzophenone) to "freeze" the interaction upon UV irradiation.
- Scaffold Diversification: We synthesize derivatives of natural products to ensure that the chemical modification does not compromise biological potency.
Our CC-ABPP Workflow
Step 1
Probe Design and Synthesis
Based on the structure of the bioactive compound, a clickable probe is rationally designed and synthesized.
Step 2
Biological Labeling
The probe is incubated with live cells, tissues, or lysates to allow interaction with active target proteins under native conditions.
Step 3
Click Chemistry Conjugation
A copper-catalyzed or copper-free click reaction is performed to attach reporter tags such as biotin or fluorescent dyes.
Step 4
Protein Enrichment and Separation
Labeled proteins are enriched using affinity purification and separated by SDS-PAGE or directly processed for proteomics.
Step 5
Mass Spectrometry Identification
High-resolution LC–MS/MS is used to identify and quantify probe-labeled proteins.
Step 6
Bioinformatics & Validation
Data is filtered against control groups (e.g., competition assays with the "cold" parent compound) to distinguish true targets from background noise.
Advantages of CC-ABPP Platform
- Function-based Profiling: Identifies active proteins rather than merely expressed proteins
- High Specificity and Sensitivity: Covalent or proximity-based labeling improves signal-to-noise ratio
- Applicable to Complex Systems: Works in cells, tissues, and in vivo models
- Broad Target Coverage: Suitable for enzymes, scaffolding proteins, and signaling regulators
- Unbiased Discovery: Enables identification of novel and unexpected targets
CC-ABPP represents a powerful and versatile technology for target identification and mechanism-of-action studies in drug discovery. Leveraging our integrated technology platform, advanced probe design capabilities, and rigorous workflows, we enable researchers to unlock the full therapeutic potential of bioactive natural products with confidence and precision.
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