Proteasome Activation (Assembly) Fluorometric Assay Kit

Additional Information

Product Name:

Also Known As:

N/A

Catalog No.:

J4000

Size:

1 Kit

Molecular Weight:

N/A

Species:

N/A

Source:

N/A

Stock:

N/A

Concentration:

See tube label

Quality Assurance:

Inhibitor and substrate > 97% by HPLC; Proteasome activity assays.

Storage:

Store at -80°C; avoid multiple freeze-thaw cycles

PDF Data Sheet:

PDF datasheet, protocol; MSDS

NCBI RefSeq:

N/A

Image(s):

J4000 Image:
(Click image to enlarge)

(A) The Mca fluorescence were recorded in reactions containing 4 nM 20S bovine proteasome, 4 nM bovine 26S proteasome, or 4 nM 20S bovine 20S proteasome + 32 nM PA700 using a plate reader with excitation and emission wavelengths at 320 and 405 nm, respectively. (B) The slope values of curves in (A) that represent the activity of each sample were plotted as a bar graph.

Shipping Method:

Wet ice shipping

References:

N/A

Details

The Proteasome Activation (Assembly) Fluorometric Assay Kit can be used for 1) in vitro assaying 20S proteasome activation (gate opening) by assembling with a regulatory particle; and 2) determining the activity of activated proteasomes in cells or tissues. The supplied Mca-KKVAPYPME-Dap(Dnp)-NH2 is sufficient for 500 X 50 µl reactions.

Description

Mca-KKVAPYPME-Dap(Dnp)–NH2 is a fluorogenic peptide substrate of proteasomes, in which the Mca fluorescence is internally quenched by Dap(Dnp). This substrate can be cleaved by mammalian, yeast or archaea activated proteasomes. Upon cleavage, the Mca fluorescence can be measured at excitation/emission wavelengths of 320/405 nm using a plate reader or fluorometer. Due to its long sequence as a nanomer, latent 20S proteasome has very weak activity on hydrolysis of Mca-KKVAPYPME-Dap(Dnp)–NH2, but it can be cleaved by activated proteasomes, including the 26S proteasome.

Components

• 1000X Mca-KKVAPYPME-Dap(Dnp)-NH2 ....................25 µl
• HEK293 cell extracts (5 mg/ml)......................................100 µl
• MG-132..........................................................................1 mg
• DMSO............................................................................200 µl
• ATP (0.5 M)....................................................................100 µl
• MgCl₂ (1 M).............................................................250 µl
• 20X Proteasome Assay Buffer........................................1.35 ml