Biotin-LLY-FMK combines a hydrophobic LLY motif with an FMK warhead and a biotin handle. The peptide targets proteases with preference for aromatic- and leucine-rich substrates. Researchers use it for affinity labeling, activity profiling, and structural studies. Its dual-function design facilitates both trapping and detection of catalytic partners.
CAT No: HB00038
Biotin-LLY-FMK is a specialized peptide-based biochemical probe that incorporates a biotin tag and a fluoromethyl ketone (FMK) reactive group at the C-terminus. This compound is designed for the targeted, irreversible labeling of proteases, particularly those recognizing the Leu-Leu-Tyr (LLY) peptide motif. The biotin moiety enables facile affinity capture and detection, while the FMK group covalently modifies active-site cysteine residues in proteases, rendering them inactive. Such structural features make Biotin-LLY-FMK a valuable reagent for chemical biology, enzymology, and proteomics research, where precise mapping and inhibition of proteolytic activity are required.
Enzyme activity profiling: Biotin-LLY-FMK serves as a powerful activity-based probe for the detection and profiling of cysteine proteases in complex biological samples. By exploiting the FMK group's ability to form a covalent bond with active protease sites, researchers can selectively label enzymes that recognize the LLY peptide sequence. The biotin tag facilitates subsequent enrichment and visualization of labeled proteases using streptavidin-based detection systems, enabling sensitive assessment of enzyme activity states in cell lysates, tissue extracts, or biochemical assays.
Target identification and validation: In drug discovery and mechanistic studies, this biotinylated FMK peptide is instrumental for identifying the molecular targets of protease inhibitors or modulators. Following incubation with biological samples, covalently modified proteins can be isolated via streptavidin affinity matrices and analyzed by mass spectrometry or immunodetection. This approach supports the elucidation of protease substrate specificity, off-target interactions, and the direct confirmation of enzyme-inhibitor engagement within complex proteomes.
Protease inhibitor screening: The compound is well-suited for use in high-throughput screening platforms aimed at discovering or characterizing novel protease inhibitors. By serving as a competitive substrate or covalent trap, it allows assessment of candidate molecules' ability to block active-site labeling. Quantifying the reduction in biotinylated enzyme signal provides a robust, mechanism-based readout for inhibitor potency and selectivity, facilitating the advancement of lead compounds in pharmaceutical and biochemical research.
Functional proteomics: Biotin-LLY-FMK is widely utilized in functional proteomics workflows to map the active protease repertoire within cells or tissues under various physiological or experimental conditions. Its application enables researchers to distinguish between inactive zymogens and catalytically competent enzymes, thereby providing insights into protease regulation, signaling pathways, and disease-associated proteolytic events. The affinity tag further supports downstream identification and characterization of labeled proteins by mass spectrometry-based proteomic techniques.
Cell-based assay development: The unique combination of peptide selectivity, irreversible inhibition, and biotin labeling makes this compound an effective tool for designing cell-based assays that monitor protease activity in situ. Researchers employ it to investigate dynamic changes in protease function in response to stimuli, genetic perturbations, or pharmacological interventions. The ability to capture and analyze active proteases directly from living cells enhances experimental resolution and supports a wide range of cellular and molecular biology applications.
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