Optineurin (D474N) [GST-tagged]

Catalogue Number
Product Size
50 µg
Price £
Accession Number
Residues Expressed
Certificate of Analysis Size
50 µg
50 µg
0.5 mg/ml
50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol
Molecular Weight
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_001008214. For full protein sequence information download the Certificate of Analysis pdf.
QA; Protein Identification
Confirmed by mass spectrometry.
QA Activity

Ubiquitin Binding Domain Activity: The lack of GST-Optineurin (D474N) ubiquitin chain binding domain activity was determined through its inability to capture polyubiquitylated IRAK1 from a lysate preparation derived from IL-1 stimulated HEK293 cells. GST-Optineurin (D474N) was pre-incubated with Glutathione Sepharose 4B for 20 minutes at 4°C followed by an incubation for 2 hours at 4°C with 2mg IL-1 stimulated HEK293 cell lysate. The binding reaction was then centrifuged and the pellet analysed by SDS-PAGE/Western blotting (Lane 2).This sample was compared alongside similarly derived pull-down from control reactions containing GST-Optineurin wild-type versus mutant (D474N) incubated in the presence of lysates derived from either IL-1 stimulated or non-stimulated HEK293 cells (Lanes 1-4). Ubiquitylated IRAK1 was identified by Western Blotting using an anti-IRAK1 antibody and such species were observed only in the pellet sample derived from a binding reaction containing wild-type GST-Optineurin and IL-1 stimulated HEK293 cell lysate (Lane 1).


Ubiquitin signals are decoded in cells by at least 200 ubiquitin binding proteins, which interact with different types of polyubiquitin chains and ubiquitin-like modifiers. These interactions induce conformational changes that allow these proteins to transmit the ubiquitin signal to effector proteins (Dikic et al., 2009). Optineurin is a protein that is most closely related to NFκB Essential Modifier (NEMO) and, like NEMO, it contains a domain that binds to both Lys63-linked and linear polyubiquitin chains (Gleason et al.,2011). These polyubiquitin chains can then regulate downstream signaling events by inducing conformational changes that activate protein kinases such as IκB kinase (IKK) or Tank binding kinase (TBK1) (Gleason et al., 2011). TBK1 can also phosphorylate optineurin at Ser177, enhancing its interaction with the microtubule-associated protein light chain 3 (LC3) which in turn promotes the autophagic clearance of ubiquitylated cytosolic Salmonella (Wild et al., 2011). Mutations in optineurin cause three different diseases in humans, namely a form of glaucoma (Rezaie et al., 2002), Paget's disease of bone (Albagha et al., 2010) and amyotrophic lateral sclerosis (ALS), a form of motor neurone disease (Maruyama et al., 2010).The Optineurin [E478G] mutation, which causes ALS, abolishes binding to polyubiquitin chains (Gleason et al., 2011). Optineurin is a powerful reagent for capturing the Lys63-linked and linear polyubiquitin chains and their binding partners present in cell extracts. It is recommended that the Optineurin [D474N] mutant, which is unable to bind polyubiquitin chains, is used as a control in such experiments (Sudhakar et al., 2009).


Albagha OM, Visconti MR, Alonso N, Langston AL, Cundy T, Dargie R, et al. (2010) Genome-wide association study identifies variants at CSF1, OPTN and TNFRSF11A as genetic risk factors for Paget's disease of bone. Nature Genetics 42, 520524.

Dikic I, Wakatsuki S and Walters KJ (2009) Ubiquitin-binding domains - from structures to functions. Nat Rev Mol Cell Biol 10, 659-671.

Gleason CE, Ordureau A, Gourlay R, Arthur JS and Cohen P (2011) Polyubiquitin binding to optineurin is required for optimal activation of TANK-binding kinase 1 and production of interferon beta. J Biol Chem 286, 35663-35674.

Maruyama H, Morino H, Ito H, Izumi Y, Kato H, Watanabe Y, et al. (2010) Mutations of optineurin in amyotrophic lateral sclerosis. Nature 465, 223-226.

Rezaie T, Child A, Hitchings R, Brice G, Miller L, Coca-Prados M, et al. (2002) Adult-onset primary open-angle glaucoma caused by mutations in optineurin. Science 295, 1077-1079.

Sudhakar C, Nagabhushana A, Jain N and Swarup G (2009) NF-kappaB mediates tumor necrosis factor alpha-induced expression of optineurin, a negative regulator of NF-kappaB. PLoS One 4, e5114.

Wild P, Farhan H, McEwan DG, Wagner S, Rogov VV, Brady NR, et al. (2011) Phosphorylation of the autophagy receptor optineurin restricts Salmonella growth. Science 333, 228-233.