Enabling Protein Degradation Drug Discovery

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  • Name
    Catalogue Number
    Size
    Price
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  • Name:
    RNF11 [6His-tagged]
    Catalogue Number:
    63-0023-025
    Size:
    25 µg
    Price:
    £100
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  • Species
    Human
  • Source
    E. coli expression
  • Quantity
    25 μg
  • Storage
    -70°C
  • Concentration
    0.5 mg/ml
  • Formulation
    50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol
  • Molecular Weight
    ~19.9 kDa
  • Stability
    12 months at -70°C; aliquot as required
  • Protein Sequence
    Accession number: NP_055187. For full protein sequence information download the Certificate of Analysis pdf.
  • QA; Protein Identification
    Confirmed by mass spectrometry.
  • QA; Activity
    E3 ligase assay: The ubiquitin conjugating activity of His-RNF11 was validated through its ability to catalyse the generation of polyubiquitin chains in the presence of the E1 activating enzyme His-UBE1, the E2 conjugating enzyme His-UBE2D1 (UbcH5a) (several E2s were tested, data generated with this E2 is provided by way of example) and ubiquitin. Incubation of His-RNF11 for 120 minutes at 37°C in the presence of ubiquitin, His-UBE1, His-UBE2D1 and ATP (Lane 1) was compared alongside two control reactions with either ATP (Lane 2) or His-RNF11 (Lane 3) excluded from the reaction. Ubiquitin conjugates were identified by Western blotting using an anti-ubiquitin conjugate antibody and these were observed only in the presence of both ATP and His-RNF11.

The enzymes of the ubiquitylation pathway play a pivotal role in a number of cellular processes including the regulated and targeted proteasome-dependent degradation of substrate proteins. Three classes of enzymes are involved in the process of ubiquitylation; activating enzymes (E1s), conjugating enzymes (E2s) and protein ligases (E3s). Ring Finger Protein 11 (RNF11) is a member of the E3 protein ligase family and cloning of the human gene was first described by Seki et al. (1999). The intrinsic E3 ligase activity of RNF11 is conferred through a RING domain at the C-terminus of the protein. RNF11 has been shown to interact with the HECT-type E3 ubiquitin ligases Nedd4, AIP4, SMURF1 and SMURF2, as well as with Cullin1, the core protein of the multi-subunit SCF E3 ubiquitin ligase complex (Santonico, Belleudi et al., 2010; Kitching et al., 2003). RNF11 has been found to mediate the ubiquitylation of AMSH by the E3 ligase SMURF2. It is thought that RNF11 recruits AMSH to SMURF2 for ubiquitylation, leading to its degradation by the 26S proteasome (Li and Seth 2004). 

References:

Kitching, R., M. J. Wong, et al. (2003) The RING-H2 protein RNF11 is differentially expressed in breast tumours and interacts with HECT-type E3 ligases. Biochim Biophys Acta 1639(2), 104-12.

Li, H. and A. Seth (2004) An RNF11: Smurf2 complex mediates ubiquitination of the AMSH protein. Oncogene 23(10), 1801-8.

Santonico, E., F. Belleudi, et al.(2010) Multiple modification and protein interaction signals drive the Ring finger protein 11 (RNF11) E3 ligase to the endosomal compartment. Oncogene 29(41), 5604-18.

Seki, N., A. Hattori, et al. (1999) Cloning and expression profile of mouse and human genes, Rnf11/RNF11, encoding a novel RING-H2 finger protein. Biochim Biophys Acta 1489(2-3), 421-7.