Specifications

The proteasome is a large, multi-catalytic protease complex within the eukaryotic cell. The 26S proteasome, a 60-subunit particle, is composed of a 20S core and one or two 19S regulatory particles. The catalytic core known as the 20S proteasome (700 kDa) is composed of two sets of seven β-subunits and two sets of seven α-subunits. The β1, β2, and β5 subunits possess caspase-like, trypsin-like, and chymotrypsin-like peptidolytic activities, respectively. In eukaryotic cells, the 20S proteasome is assembled with the aid of at least five assembly chaperones. The degradation of cellular proteins by the proteasome is initiated by attachment of an ubiquitin chain to the polypeptide. The ubiquitin tag is recognized by and binds to the ‘lid’, a 19S particle, followed by disassembly of ubiquitin chains, protein unfolding and subsequent translocation into the 20S proteasome.

Proteasomal activity plays essential role in major cellular events including protein quality control, regulation of gene transcription, DNA damage response, aging, oxidative stress and signal transduction. Many selective proteasome inhibitors targeting specific active sites have been identified in the last two decades. Most notably, Bortezomib and Carfilzomib are FDA-approved proteasome inhibitor drugs for the treatment of multiple myeloma. Several new proteasome inhibitors are currently in trials for the treatment of multiple myeloma and other malignancies.

This purified 20S proteasome preparation can be used in vitro for the degradation of peptide substrates and unfolded proteins. Chymotrypsin-like activity of the proteasome is readily measured using Suc-LLVY-AMC (Cat. # PS500). Upon cleavage by the active enzyme, it generates a highly fluorescent product with an emission wavelength at 460 nm.

Note:

• The chymotrypsin-like activity of the 20S proteasome (45µg/ml) was measured using 100 µM Suc-LLVY-AMC in 20 mM HEPES, pH 7.5, 0.5 mM EDTA, 0.05% Triton X, plus or minus 0.035% SDS.

• The chymotrypsin-like activity of the 20S proteasome (18 µg/ml) was measured using 100 µM Suc-LLVY-AMC in 20 mM HEPEs, pH 7.5, 0.5 mM EDTA, 0.05% Triton X, plus or minus 0.5 mM MG132 (Cat. # SI9710).

Info

Source	Human red blood cells
Quantity	50 µg at 0.5 mg/mL
State	Liquid (buffer: 20 mM Tris-HCl pH 7.2, 10% Glycerol, 150 mM KCl, 1 mM β– mercaptoethanol)
Storage	-80°C; Avoid repeated freeze/thaw cycles

Life Sensors的泛素和泛素链泛素是一种小的多肽，通过其C端与目标蛋白上的赖氨酸的ε-氨基缀合。这种缀合称为单泛素化。随后，可以利用遍在蛋白表面上的七个赖氨酸残基（K6，K11，K27，K29，K33，K48，K63）中的任何一个，将额外的遍在蛋白部分与初始遍在蛋白结合。这种泛素链形成称为多泛素化。泛素的酶结合是通过一系列酶进行的，即泛素激活酶E1，泛素结合酶E2和泛素连接酶E3。蛋白质的泛素化在细胞中是可逆的。单泛素化和多泛素化的链均被去泛素化酶催化的水解作用裂解 （DUB）。LifeSensors在遍在泛素途径的所有水平的酶（包括E1，E2和E3酶和DUB）上都有丰富的经验。LifeSensors对蛋白质纯化的广泛了解使我们能够生产大约30种E2酶，20种E3酶和35个DUB，并具有定制表达的能力。此外，我们广泛的检测方法简化了确认酶活性和确定E2-E3对兼容性的过程，并确保您可以选择最能解决希望探索的特定，关键生物学问题的酶。LifeSensors已为您的研究开发了多种泛素衍生物，包括用于结合相互作用研究的DUB耐药性聚泛素链，用于DUB分析和筛选的泛素结合物（荧光团和报告酶），选择性泛素链（二，三和四环素）泛素链）用于DUB特异性和结合测定，位点特异性赖氨酸泛素突变体用于E3连接酶和DUB选择性测定。