His6-NEDP1, human recombinant

Catalog number: SBB-DE0025, 50μg

NEDP1 is a protease which hydrolyzes the isopeptide bond between the C-terminus of NEDD8 and the respective lysine of the substrate protein to which the NEDD8 is covalently bound. This NEDP1 is his-tagged on the N-terminus, recombinantly expressed in E. coli.[1][2]

In stock
SKU
0025
$149.00

Description

NEDP1 (NEDD8-specific protease 1)-is a deconjugating enzyme with specificity towards NEDDylated substrates. NEDP1 discriminates between ubiquitinated and NEDDylated targets, despite similar C-terminal regions. NEDP1 is important in regulating cullin associated ubiqutin ligase enzymes such as those of the SCF(Skp-Cullin-Fbox) family and subsequently, it’s important in regulating cell cycle progression and other biological functions. This NEDP1 is recombinantly expressed in E.coli, and contains an N-terminal His6-tag.

For Research Use Only, Not For Use In Humans.

Specifications

Product Overview
Quantity: 50 μg
Molecular Weight: 26.5 kDa
Purity: >95% by SDS-PAGE
Substrate Properties: His6-tagged, N-terminus. Working concentrations of this enzyme range from 1 to 10 nM.
Storage Buffer: 50 mM HEPES pH 7.5, 100 mM NaCl, 1 mM TCEP
Storage Store at −80°C after product arrival. Avoid multiple freeze / thaws. It is recommended to make multiple aliquots after the first thaw.

Figures & Data

His6-NEDP1, human recombinant

Figure 1. His6-NEDP1, SDS-PAGE From left to right, increasing amounts of ubiquitin loaded onto a 4-12 % SDS-PAGE gel, stained with Coomassie blue.

His6-NEDP1 Michaelis–Menten Kinetics

Figure 2. Michaelis–Menten Kinetics NEDD8 Rhodamine 110 serially diluted from 1.6 to 0.0125 uM was digested with 30pM NEDP1 over time. The assay was carried out in a reaction buffer of 50 mM HEPES pH 7.5, 1 mM TCEP, 0.1 mg/ml BSA, at 25C. Initial velocities at each substrate concentration were plotted and fit to the Michaelis–Menten equation. Kinetic parameters were calculated at: Km= 0.087 μM, Vmax = 4.44 nMs-1, kcat= 148s-1, kcat/Km= 1.70 x 109 M-1s-1.

Citations & References

1) Mendoza, Heidi M., et al. "NEDP1, a highly conserved cysteine protease that deNEDDylates Cullins." Journal of Biological Chemistry 278.28 (2003): 25637-25643

2) Shen, Lin‐nan, et al. "Structural basis of NEDD8 ubiquitin discrimination by the deNEDDylating enzyme NEDP1." The EMBO journal 24.7 (2005): 1341-1351