PolySUMO-Qapture kit



BML-UW0955-0001	1 Kit	358.00 USD

Figure: Western blot analysis of polySUMO-Qapture enrichment of lysate derived polySUMO modified proteins. PolySUMO-Qapture experiment set-up and run as described in the manual in “Capture / enrichment of polySUMOylated proteins”. PolySUMO-conjugates present in Starting Material, Unbound Fraction and Elution Fraction were detected by Western blotting as described in “Analysis by Western blotting”, using the provided SUMO-2/3 (Prod. No. BML-UW0510) antibody at a dilution of 1:500. Capture of SUMO-protein conjugates from Control SUMOylated-protein lysate (Prod. No. BML-UW0985). Key: SM = Starting Material, UF = Unbound Fraction and EL = Elution Fraction.

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Product Specification

Application:	For the isolation and enrichment of polySUMOylated proteins. 1. Capture and isolation of polySUMOylated protein conjugates from cell/tissue lysates of interest with subsequent detection and analysis by Western blotting. 2. Biochemical characterisation, secondary purification by substrate-specific immunoprecipitation, or identification of polySUMO modified proteins by proteomic analysis following release of free polySUMOylated proteins in their active/native form by specific elution from the affinity matrix,. 3. Selective purification/pull down of polySUMOylated proteins from in vitro SUMOylation assays.

Quantity:	Sufficient for 10 binding assays.

Use/Stability:	PolySUMO-Qapture-T matrix should be stored at 4ºC upon receipt. All other components should be stored at –80ºC to ensure stability and activity.

Handling:	Avoid freeze/thaw cycles.

Short Term Storage:	-80°C

Long Term Storage:	-80°C

Kit/Set Contains:	PolySUMO-Qapture matrix (Prod. No. BML-UW0995) 200µL settled resin provided, 50% suspension. Control SUMOylated-protein lysate (Prod. No. BML-UW0985) 220 µg (110 µL), 2mg/mL in 50mM TRIS-Cl, pH 7.5, 150mM sodium chloride, 1% NP-40, 0.5% deoxycholate, 200mM iodoacetamide. PolySUMO-Qapture elution buffer (Prod. No. BML-UW1005) 1mL. SUMO-2/3, sheep polyclonal antibody (Prod. No. BML-KW0510) 10µL. 0.5mL Screw capped tube pack (Prod. No. BML-UW0990) 10 Tubes.

Miscellaneous/General:	The PolySUMO-Qapture kit is an efficient tool for the selective isolation of polySUMOylated proteins. The kit facilitates the affinity purification of polySUMOylated proteins from cell extracts and tissue lysates using a high-binding multi-SIM-containing affinity matrix, whilst discriminating against free and mono-SUMO-conjugate binding. Captured proteins are specifically eluted prior to analysis by Western blotting, using the SUMO2/3 antibody provided or antibodies to specific proteins of interest, further purification by substrate specific immunoprecipitation, biochemical characterization, or potential substrate identification by proteomic methods. PolySUMO-conjugate-containing samples are prepared in native form in the presence of protease inhibitors to prevent loss through the action of deSUMOulating enzymes. Optimisation of binding permits complete isolation of the full range of polySUMO-protein conjugates from a specific lysate. The kit is also supplied with a high quality polySUMOylated protein solution for use as a positive control in polySUMO-Qapture assays.

General Literature References

PARP-1 transcriptional activity is regulated by sumoylation upon heat shock: N. Martin, et al.; EMBO J. 28, 3534 (2009), Abstract;

System-wide changes to SUMO modifications in response to heat shock: F. Golebiowski, et al.; Sci. Signal. 2, ra24 (2009), Abstract;

Architecture and assembly of poly-SUMO chains on PCNA in Saccharomyces cerevisiae: H. Windecker & H.D. Ulrich; J. Mol. Biol. 376, 221 (2008), Abstract;

In vivo identification of human small ubiquitin-like modifier polymerization sites by high accuracy mass spectrometry and an in vitro to in vivo strategy: I. Matic, et al.; Mol. Cell Proteomics 7, 132 (2008), Abstract;

In vivo modeling of polysumoylation uncovers targeting of Topoisomerase II to the nucleolus via optimal level of SUMO modification: Y. Takahashi & A. Strunnikov; Chromosoma 117, 189 (2008), Abstract;

RNF4 is a poly-SUMO-specific E3 ubiquitin ligase required for arsenic-induced PML degradation: M.H. Tatham, et al.; Nat. Cell Biol. 10, 538 (2008), Abstract;

The fast-growing business of SUMO chains: H.D. Ulrich; Mol. Cell. 32, 301 (2008), Abstract;

The Ulp2 SUMO protease is required for cell division following termination of the DNA damage checkpoint: D.C. Schwartz, et al.; Mol. Cell Biol. 27, 6948 (2007), Abstract;

Specification of SUMO1- and SUMO2-interacting motifs: C.M. Hecker, et al.; J. Biol. Chem. 281, 16117 (2006), Abstract;

SUMO modifications control assembly of synaptonemal complex and polycomplex in meiosis of Saccharomyces cerevisiae: C.H. Cheng, et al.; Genes Dev. 20, 2067 (2006), Abstract;

Polymeric chains of SUMO-2 and SUMO-3 are conjugated to protein substrates by SAE1/SAE2 and Ubc9: M.H. Tatham, et al.; J. Biol. Chem. 276, 35368 (2001), Abstract;

SUMO, ubiquitin’s mysterious cousin: S. Muller, et al.; Nat. Rev. Mol. Cell Biol. 2, 202 (2001), Abstract;

Functional heterogeneity of small ubiquitin-related protein modifiers SUMO-1 versus SUMO-2/3: H. Saitoh & J. Hinchey; J. Biol. Chem. 275, 6252 (2000), Abstract;