Suc-Leu-Leu-Val-Tyr-AMC

Suc-Leu-Leu-Val-Tyr-AMC is a fluorogenic substrate. Sequence: Suc-Leu-Leu-Val-Tyr.

Signaling Pathways >> Others >> Others

Dye Reagents

Peptides

Research Areas >> Others

Suc-Leu-Leu-Val-Tyr-AMC (Suc-LLVY) is a membrane-permeable calpain-specific fluorogenic substrate, pteolytic hydrolysis of the peptidyl-7-amino bond liberates the highly fluorescent 7-amino-4-methylcoumarin (AMC) moiety[1]. The effectof TGF-β on hydrolysis of these substrates (e.g Suc-Leu-Leu-Val-Tyr-AMC) are assessed. Biliary epithelial H69 cells are incubated with 10, 1, 0.1, or 0 ng/mL TGF-β for 24 h. Substrate hydrolysis is then fluorometrically assessed in cytosolic extracts. Basal activity is 1.12, 8.33, and 14.52 nmol AMC/mg protein/min for suc-LLVY-AMC, z-LLE-AMC, and z-LLL-AMC hydrolysis, respectively[2].

Immunoprecipitation is carried out for the two sets of samples, using the same amount of protein. The 20 S and 26 S proteasome immunoprecipitates are washed with 50 mM Hepes/KOH (pH 7.5), and 50 mM Hepes/KOH (pH 7.5) containing 2 mMATP, respectively, prior to the determination of peptidase activity using 50 μM suc-Leu- Leu-Val-Tyr-AMC as substrate in these buffers[3].

[1]. Roberta L. Debiasi, et al. Reovirus-Induced Apoptosis Is Preceded by Increased Cellular Calpain Activity and Is Blocked by Calpain Inhibitors. J Virol. 1999 Jan; 73(1): 695–701.

[2]. Tadlock L, et al. Transforming growth factor-beta inhibition of proteasomal activity: a potential mechanism of growth arrest. Am J Physiol Cell Physiol. 2003 Aug;285(2):C277-85. Epub 2003 Mar 19.

[3]. Gardner RC, et al. Characterization of peptidyl boronic acid inhibitors of mammalian 20 S and 26 S proteasomes and their inhibition of proteasomes in cultured cells. Biochem J. 2000 Mar, 2:447-54.

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