TAME

TAME is an inhibitor of anaphase-promoting complex (APC), which prevents its activation by Cdc20 and Cdh1; TAME is also an inhibitor of cyclin proteolysis in mitotic Xenopus egg extract, with an IC50 of 12 µM.

Signaling Pathways >> Cell Cycle/DNA Damage >> APC

Research Areas >> Cancer

APC

TAME (tosyl-L-arginine methyl ester, 200 μM) shows inhibitory effect on APC activation when added to mitotic Xenopus extract. TAME (1-200 μM) inhibits Cdc20 association with mitotic APC and also inhibits Cdh1 association with interphase APC. TAME significantly inhibits crosslinking of the IR peptide to Cdc27 and Cdc16 but only slightly reduces crosslinking to Cdc23 and Apc7 at 20 µM; TAME strongly inhibits crosslinking to all APC subunits at 200 µM. TAME (200 µM) also inhibits binding of wild type Cdc20 to the APC, but not binding of a ΔIR mutant[1]. TAME (200 μM) induces Cdc20 dissociation from the APC in mitotic Xenopus extract calls for APC-dependent ubiquitination. TAME induces Cdc20 dissociation from the APC by promoting Cdc20 ubiquitination, but the dissociation can be suppresses by Cyclin B1. TAME terminates cyclin B1 ubiquitination prematurely[2]. TAME interacts with ATP by β and γ phosphate and the adenine ring of ATP. TAME in combination with the Mg(II) ion accelerates the ATP hydrolysis process[3].

APC is first immunoprecipitated from mitotic Xenopus extract and 200 μM TAME is added to the extract for 15 min to drive Cdc20 dissociation. The beads are then washed quickly with XB + 0.1% IGEPAL CA-630 4 times and then washed with XB + 0.1% IGEPAL CA-630 for 5 min × 2 while kept shaking vigorously[2].

[1]. Zeng X, et al. Pharmacologic inhibition of the anaphase-promoting complex induces a spindle checkpoint-dependent mitotic arrest in the absence of spindle damage. Cancer Cell. 2010 Oct 19;18(4):382-95.

[2]. Zeng X, et al. An APC/C inhibitor stabilizes cyclin B1 by prematurely terminating ubiquitination. Nat Chem Biol. 2012 Feb 26;8(4):383-92.

[3]. Ma Y, et al. Differential effects of Mg(ii) and N(alpha)-4-tosyl-l-arginine methyl ester hydrochloride on the recognition and catalysis in ATP hydrolysis. Dalton Trans. 2008 Feb 28;(8):1081-6.

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