Ac-Cys(dodecyl)-chloromethylketone is a cysteine-derived amino acid derivative in which the thiol side chain is modified with a dodecyl substituent and the N-terminus is acetylated (Ac-), yielding a thioether-bearing, protected-like cysteine analogue. The molecule contains an amide linkage from the acetyl group, a carboxyl-derived functionality consistent with a cysteine scaffold, and a chloromethyl ketone electrophile (-CH2Cl-C(=O)-) that provides a reactive carbonyl center for covalent capture by nucleophiles in chemical biology workflows. It is used as a substrate-mimicking or labeling reagent in studies of protease active-site chemistry and in the preparation of more complex cysteine-functionalized peptide or small-molecule derivatives via thiol-directed conjugation chemistry.
CAT No: CP26884
CAS No:253589-60-5
Synonyms/Alias:Acetyl-Cys(dodecyl)chloromethylketone;253589-60-5;CHEMBL540205;CTK8E7843;AC-CYS-CHLOROMETHYLKETONE;ZINC14096586;RT-011181;N-[(R)-1-(Dodecylthiomethyl)-2-oxo-3-chloropropyl]acetamide
Ac-Cys(dodecyl)-chloromethylketone is a cysteine-targeting chloromethyl ketone derivative used as a covalent inhibitor scaffold in chemical biology and protease research. The molecule incorporates an N-terminal acetylated cysteine framework and a dodecyl side chain, providing a hydrophobic handle that supports membrane-proximal or lipophilic microenvironment studies. As a reactive chloromethyl ketone electrophile, it is commonly employed to form irreversible adducts with active-site cysteine residues in target proteins, enabling activity-based profiling and functional interrogation.
1. Protease Activity Probing
Ac-Cys(dodecyl)-chloromethylketone is used by chemical biology groups to covalently label and inactivate active-site cysteine proteases in lysates and purified enzyme systems. Researchers rely on the chloromethyl ketone electrophile to capture catalytically competent enzyme populations, while the N-acetylated cysteine motif and dodecyl substituent help tune hydrophobic association during incubation. This format is frequently selected for activity-based studies where readouts such as gel-based detection, immunoblotting of labeled targets, or downstream enrichment workflows are used to compare enzyme activity states across conditions.
2. Activity-Based Chemical Proteomics
Ac-Cys(dodecyl)-chloromethylketone supports activity-based profiling strategies in proteomics workflows aimed at mapping cysteine protease activity in complex biological samples. In these studies, the irreversible covalent reaction with active-site cysteines enables researchers to distinguish functional enzymes from inactive or inhibited forms, improving interpretability of proteome-wide activity patterns. The hydrophobic dodecyl group can influence labeling distribution and accessibility in lipid-rich or compartmentalized samples, making the reagent a practical choice for experiments that compare activity across different sample preparations, stimulation conditions, or inhibitor panels.
3. Covalent Inhibitor Lead Studies
Ac-Cys(dodecyl)-chloromethylketone is also applied in medicinal chemistry and inhibitor discovery programs as a covalent warhead scaffold for studying cysteine-directed inhibition mechanisms in vitro. Teams developing covalent inhibitors use this type of chloromethyl ketone functionality to interrogate how electrophile reactivity and side-chain hydrophobicity affect target engagement and labeling outcomes. The N-terminal acetylated cysteine framework provides a consistent reactive context for comparing structure-activity relationships across related electrophiles, supporting early-stage optimization of covalent binding behavior in enzymology assays and cell-lysate functional screens.
4. Target Engagement and Mechanism Validation
Ac-Cys(dodecyl)-chloromethylketone is employed in target engagement experiments to validate whether a cysteine protease of interest is being modified under defined experimental conditions. Protein scientists often use covalent labeling readouts to confirm active-site accessibility and to differentiate competitive inhibition from irreversible inactivation, particularly when multiple protease families are present. The reagent's ability to form stable covalent adducts with active-site cysteine residues makes it useful for mechanism validation studies where follow-up analyses such as activity readouts, labeled protein detection, or comparative band-shift/enrichment approaches are used to interpret inhibitor effects.
1. SERS spectrum of the peptide thymosin‐β4 obtained with Ag nanorod substrate
3. Autoinhibition and phosphorylation-induced activation of phospholipase C-γ isozymes
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