SUMO antibody

Antigen: SUMO

Protein Type: Recombinant

Clonality: Polyclonal

Host: Rabbit

Reactivity: Yeast

Application: ELISA, Western Blotting (WB), Immunoprecipitation (IP)

Catalog no.: ABIN107153

Additional Information

Gene ID: 852122

UniProt: Q06

Immunogen: This purified antibody was prepared from rabbit serum after repeated immunizations with recombinant yeast SUMO protein.

Format: Lyophilized

Reconstitution: Restore with deionized water (or equivalent)

Isotype: IgG

Description: Anti SUMO Antibody recognizes SUMO. Covalent modification of cellular proteins by the ubiquitin-like modifier SUMO (small ubiquitin-like modifier) regulates various cellular processes, such as nuclear transport, signal transduction, stress responses and cell cycle progression. But, in contrast to ubiquination, sumoylation does not tag proteins for degradation by the 26S proteasome, but rather seems to enhance stability or modulate their subcellular compartmentalization. Ubiquitin-like proteins fall into two classes: the first class, ubiquitin-like modifiers (UBLs) function as modifiers in a manner analogous to that of ubiquitin. Examples of UBLs are SUMO, Rub1 (also called Nedd8), Apg8 and Apg12. Proteins of the second class include parkin, RAD23 and DSK2, are designated ubiquitin-domain proteins (UDPs). These proteins contain domains that are related to ubiquitin but are otherwise unrelated to each In contrast to UBLs, UDPs are not conjugated to other proteins. Once covalently attached to cellular targets, SUMO regulates protein:protein and protein:DNA interactions, as well as localization and stability of the target protein. Sumoylation occurs in most eukaryotic systems, and SUMO is highly conserved from yeast to humans. Where invertebrates have only a single SUMO gene termed SMT3, three members of the SUMO family have been identified in vertebrates: SUMO-1 and the close homologues SUMO-2 and SUMO-3. SUMO has been called SMT3 (yeast), sentrin, PIC1, GMP1 and UBL1. SUMO has been shown to bind and regulate mammalian SP-RINGs (such as Mdm2, PIAS and PML), RanGAP1, RanBP2, p53, p73, HIPK2, TEL, c-Jun, Fas, Daxx, TNFRI, Topo-I, Topo-II, WRN, Sp100, IkB-a, Androgen receptor (AR), GLUT1/4, Drosophila Ttk69, Dorsal, CaMK, yeast Septins, and viral CMV-IE1/2, EBV-BZLF1, HPV/BPV-E1. These bindings implicate SUMO in the stabilization of the target proteins and/or their localization to subcellular complexes. SUMO has an apparent molecular weight of ~12kDa and human SUMO-1 (a 101 amino acid polypeptide) shares 50% sequence identity with SUMO-2 and SUMO-3 and with yeast SMT3. SUMO and ubiquitin only show about 18% homology, but both possess a common three-dimensional structure characterized by a tightly packed globular fold with b-sheets wrapped around an a-helix.

Application Details

Application Notes: This purified polyclonal antibody reacts yeast SUMO by western blot and ELISA. Although not tested, this antibody is likely functional in immunohistochemistry and immunoprecipitation. For immunoblotting a 1:1,000 dilution is recommended. A 12 kDa band corresponding to yeast SUMO is detected. Most yeast cell lysates can be used as a positive control without induction or stimulation. For ELISA a 1:1,000 to 1:5,000 dilution is recommended. Researchers should determine optimal titers for other applications. For immunoblotting a 1:1,000 dilution is recommended. A 12 kDa band corresponding to yeast SUMO is detected. Most yeast cell lysates can be used as a positive control without induction or stimulation. For ELISA a 1:1,000 to 1:5,000 dilution is recommended. Researchers should determine optimal titers for other applications. For immunoblotting a 1:1,000 dilution is recommended. A 12 kDa band corresponding to yeast SUMO is detected. Most yeast cell lysates can be used as a positive control without induction or stimulation. For ELISA a 1:1,000 to 1:5,000 dilution is recommended. Researchers should determine optimal titers for other applications.
ELISA Dilution: 1:5,000 - 1:25,000
Western Blot Dilution: 1:500 - 1:3,000

Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2

Preservative: 0.01% (w/v) Sodium azide

Storage: Expiration date is one (1) year from date of opening. None. Store vial at 4° C prior to restoration. For extended storage aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4° C as an undiluted liquid. Dilute only prior to immediate use. 

Research Area: Signaling, Proteases, DNA/RNA

Restrictions: For Research Use only

Images

Anti-SUMO antibody, generated by immunization with recombinant yeast SUMO, was tested by western blot against several constructs of SUMO-GFP fusion proteins after cleavage by proteases in insect cell protein extracts. These constructs contained various linkers between the SUMO and GFP portion of the fusion proteins. Each sample was run twice. The left lanes each contain 2 ug E.coli expressed and purified SUMO-GFP fusion proteins after incubation with lysed cells (50 ug total protein) for 1 h. The right lanes contain the same fusion proteins incubated with the lysate in the presence of 2% SDS. After probing with anti-GFP antibodies the membranes were stripped of antibody using SDS-DTT solution for 30 m at 60° C and were then re-probed using the anti-SUMO antibody at a 1:1000 dilution incubated overnight at 4° C in 5% non-fat dry milk in TTBS. Detection occurred using a 1:2000 dilution of HRP-labeled Donkey anti-Rabbit IgG (code # 611-703-127) for 1 hour at room temperature. A chemiluminescence system was used for signal detection (Roche). Other detection systems will yield similar results.

Publications

Chen, Ding, LeJeune et al.: "Rpb1 sumoylation in response to UV radiation or transcriptional impairment in yeast." in: PLoS ONE, Vol. 4, Issue 4, pp. e5267, 2009 (PubMed).

Trujillo, Tyler, Ye et al.: "A genetic and molecular toolbox for analyzing histone ubiquitylation and sumoylation in yeast." in: Methods (San Diego, Calif.), 2011 (PubMed).