Desthiobiotin-iodoacetamide DbGGI

DbGGI is a bifunctional, cysteine-reactive ABPP probe that links an iodoacetamide warhead to a desthiobiotin tag via a flexible 10-atom chain, providing a "click-free" alternative to alkyne-tagged probes. This design enables strong, reversible capture on streptavidin beads and efficient peptide elution under mild acidic conditions, which eliminates background from copper reagents and yields a cleaner, more accurate representation of pulldown results for mass spectrometry or western blot. The probe′s principal innovation is its application in a quantitative, concentration-dependent enrichment strategy that selectively identifies hyper-reactive primary protein targets (PPTs) in cell lysates—a class often obscured in conventional high-concentration ABPP. As demonstrated in U87MG cell lysates, this approach captured 116 targets at a low concentration (10 µM) versus 193 at a high concentration (250 µM). Note: Differentially label cell lysates (2 mg/mL) with low (10 µM) and high (250 µM) DbGGI from a DMSO stock for 1 hour at 37°C. Then, capture the proteomic targets via a 4-hour incubation with streptavidin beads at room temperature.