While fluorimetric assay kits for the detection and inhibitor screening
of Tumor Necrosis Factor alpha (TNF-α) Converting Enzyme (TACE) currently
exist, none of these have been able to take full advantage of the performance
provided by long wavelength FRET technologies.
AnaSpec’s SensoLyte® 520 TACE Assay Kit offers
fluorescence detection at the industry’s longest wavelengths. With superior
fluorescence quantum yield and longer emission wavelength, this kit shows less
autofluorescence interference from test compounds and cellular components, and
provides better assay sensitivity than comparable kits using shorter-wavelength
520 TACE Activity Assay Kit uses a FRET peptide substrate for the
continuous measurement of enzyme activity. The FRET pair is composed of 5-FAM
(5-carboxyfluorescein) and QXL™ 520, one of AnaSpec’s proprietary fluorescence
quenchers. In the intact FRET peptide, the fluorescence of 5-FAM is quenched
by QXL™ 520. Upon cleavage of the FRET peptide by the active enzyme, the fluorescence
of 5-FAM is recovered, and can be continuously monitored at excitation/emission
= 490 nm/520 nm. The kit can be used to detect the activity of TACE or screen
for TACE inhibitors.
Tumor Necrosis Factor alpha (TNF-α) Converting Enzyme (TACE),
also called ADAM17 or α-Secretase belongs to the ADAM (A Disintegrin and
Metalloprotease) family of proteins involved in myogenesis, neurogenesis, fertilization
through the ectodomain shedding of cell surface proteins.1 TACE,
the protease responsible for the generation of mature TNF-α, plays an important
role in acute and chronic inflammation. Since TNF-α is a crucial mediator
in the inflammatory process, considerable efforts have been made in the research
and development of anti-TNF-α agents, for the purpose of reducing the severity
of inflammatory responses in disease states.2,3 The inhibition of
TACE by a pharmacological agent may represent an alternative approach to modulate
the effect of TNF-α.4 TACE is also responsible for the proteolytic
cleavage of amyloid precursor protein, L-selectin, transforming growth factor-α.1,5,6
Figure 1. TAPI-0 inhibition of TACE activity measured with
SensoLyte®520 TACE Activity Assay Kit
Looking for custom development of fluorimetric assays? Please
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email firstname.lastname@example.org to see how
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1. Peschon, JJ. et al. Science 282, 1281 (1998).
2. Feldman, M. et al. Transplant Proc. 30, 4126 (1998).
3. Siegel, SA. et al. Cytokine 7, 15 (1995)
4. Levin, JI. et al. Bioorg. Med. Chem. Lett. 13, 2799 (2003).
5. Buxbaum, JD. et al. J. Biol. Chem. 273, 27765 (1998).
6. Smalley, DM. and KJ. Ley, Cell. Mol. Med. 9, 255 (2005).