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5562
PTMScan® Ubiquitin Remnant Motif (K-ε-GG) Kit
Proteomic Analysis Products
PTMScan

PTMScan® Ubiquitin Remnant Motif (K-ε-GG) Kit #5562

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PTMScan® Ubiquitin Remnant Motif (K-ε-GG) Kit: Image 1

Motif analysis was done using all ubiquitin remnant peptides in a ubiquitin remnant motif antibody PTMScan® study. Cellular tryptic peptides from HCT116 cells. untreated or treated with MG132, were immunoprecipitated with ubiquitin remnant motif antibody K-ε-GG and analyzed by OrbiTrap MS. The study gave 599 non-redundant sites. The motif logo shows that the K-ε-GG antibody is a general motif antibody that recognizes the K-ε-GG motif and does not have other amino acid preferences.

Product Includes Volume (with Count)
PTMScan® Ubiquitin Branch Motif (K-ε-GG) Immunoaffinity Beads 10 x 80 µl
PTMScan® IAP Buffer (10X) 9993 10 x 600 µl
PTMScan® Limited Use License 1 x 1 ml

Product Usage Information

Cells are lysed in a urea-containing buffer, cellular proteins are digested by proteases, and the resulting peptides are purified by reversed-phase, solid-phase extraction. Peptides are then subjected to immunoaffinity purification using a PTMScan® Motif antibody conjugated to protein A agarose beads. Unbound peptides are removed through washing, and the captured PTM-containing peptides are eluted with dilute acid. Reversed-phase purification is performed on microtips to desalt and separate peptides from antibody prior to concentrating the enriched peptides for LC-MS/MS analysis. CST recommends the use of PTMScan® IAP Buffer #9993 included in the kit. A detailed protocol and Limited Use License allowing the use of the patented PTMScan® method is included with the kit.

Storage

Antibody beads supplied in IAP buffer containing 50% glycerol. Store at -20°C. Do not aliquot the antibody.

Product Description

PTMScan® Technology employs a proprietary methodology from Cell Signaling Technology (CST) for peptide enrichment by immunoprecipitation using a specific bead-conjugated antibody in conjunction with liquid chromatography (LC) tandem mass spectrometry (MS/MS) for quantitative profiling of post-translational modification (PTM) sites in cellular proteins. These include phosphorylation (PhosphoScan®), ubiquitination (UbiScan®), acetylation (AcetylScan®), and methylation (MethylScan®), among others. PTMScan® Technology enables researchers to isolate, identify, and quantitate large numbers of post-translationally modified cellular peptides with a high degree of specificity and sensitivity, providing a global overview of PTMs in cell and tissue samples without preconceived biases about where these modified sites occur (1). For more information on PTMScan® Proteomics Services, please visit www.cellsignal.com/common/content/content.jsp?id=ptmscan-services.

Background

Ubiquitin is a conserved polypeptide unit that plays an important role in the ubiquitin-proteasome pathway. Ubiquitin can be covalently linked to many cellular proteins by the ubiquitination process, which targets proteins for degradation by the 26S proteasome. Three components are involved in the target protein-ubiquitin conjugation process. Ubiquitin is first activated by forming a thiolester complex with the activation component E1; the activated ubiquitin is subsequently transferred to the ubiquitin-carrier protein E2, then from E2 to ubiquitin ligase E3 for final delivery to the epsilon-NH2 of the target protein lysine residue (2-4). The ubiquitin-proteasome pathway has been implicated in a wide range of normal biological processes and in disease-related abnormalities. Several proteins such as IκB, p53, cdc25a and bcl-2 have been shown to be targets for the ubiquitin-proteasome process as part of regulation of cell cycle progression, differentiation, cell stress response, and apoptosis (5-8).

In this assay, the PTMScan® Ubiquitin Remnant Motif (K-ε-GG) Antibody Bead Conjugate, a proprietary ubiquitin branch (“K-ε-GG”) antibody with specificity for a di-glycine tag that is the remnant of ubiqui- tin left on protein substrates after trypsin digestion, is used to enrich ubiquitinated peptides from trypsin digested cell samples. This enrichment is followed by LC-MS/MS analy- sis for quantitative profiles of hundreds to over a thousand non-redundant ubiquitinated sequences.

  1. Rush, J. et al. (2005) Nat Biotechnol 23, 94-101.
  2. Ciechanover, A. (1998) EMBO J 17, 7151-60.
  3. Hochstrasser, M. (2000) Nat Cell Biol 2, E153-7.
  4. Hochstrasser, M. (2000) Science 289, 563-4.
  5. Bernardi, R. et al. (2000) Oncogene 19, 2447-54.
  6. Aberle, H. et al. (1997) EMBO J 16, 3797-804.
  7. Salomoni, P. and Pandolfi, P.P. (2002) Nat Cell Biol 4, E152-3.
  8. Jesenberger, V. and Jentsch, S. (2002) Nat Rev Mol Cell Biol 3, 112-21.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST's products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST's Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

For Research Use Only. Not For Use In Diagnostic Procedures.
AcetylScan is a trademark of Cell Signaling Technology, Inc.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
MethylScan is a trademark of Cell Signaling Technology, Inc.
PTMScan is a trademark of Cell Signaling Technology, Inc.
UbiScan is a trademark of Cell Signaling Technology, Inc.
Use of Cell Signaling Technology (CST) Motif Antibodies within certain methods (e.g., U.S. Patents No. 7,198,896 and 7,300,753) may require a license from CST. For information regarding academic licensing terms please have your technology transfer office contact CST Legal Department at [email protected] For information regarding commercial licensing terms please contact CST Pharma Services Department at [email protected]