Specifications

“My laboratory has published two recent manuscripts reporting the use of FLAG-K63 and FLAG K48 TUBE reagents. We used these reagents to study the ubiquitination of the nonstructural 2 transmembrane protein of hepatitis C virus (HCV) by the cellular E3 ligase MARCH8. By being quite selective and easy to handle, these reagents helped us characterize the specific ubiquitin modification.”

Dr. Shirit Einav, Associate Professor, Department of Microbiology and Immunology, Stanford University of Medicine

FLAG® Anti-K63 TUBE is an ideal reagent for efficient isolation and enrichment of K63-polyubiquitinated proteins from cell & tissue extracts or in vitro synthesized mixtures. Based on a peptide engineered by Sims et al., it consists of multiple ubiquitin interaction motifs (UIMs) joined by a rigid, helical linker that spaces the UIMs for selective binding to extended K63-linked polyubiquitin chains. The result is a peptide the exhibits high affinity binding to K63-linked polyubiquitin together with 1000 to 10,000-fold selectivity over K48- and K11- linkages. Expression of this peptide in vitro inhibits K63-linked polyubiquitin-dependent processes and protects K63-linked polyubiquitin chains from degradation. Combining this peptide with a FLAG epitope tag generates, for the first time, a powerful affinity reagent suitable for isolation, purification and characterization of proteins modified by K63-linked polyubiquitin. Flag K63-TUBE 1 allows isolation of K63-linked polyubiquitin without the need for overexpression of ubiquitin mutants, tagged ubiquitins or the inclusion of DUB inhibitors any of which could alter cellular physiology. Isolated ubiquitinated proteins can be characterized by Western blot, mass spec or further biochemical analysis. (FLAG is a registered trademark of Sigma-Aldrich Corporation LLC).

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Info

Molecular Weight	9062.7 (calculated)
Formulation	1 mg/mL (110uM) in PBS, pH 7.2
Recommended Concentration	50-500 nM

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。