AQUA Custom Services: Quantifying Poly-Ubiquitin Chains

Ubiquitination is a post-translational modification that plays an important role in nearly every cellular process. Target proteins may be modified with Ubiquitin in a variety of ways including poly-ubiquitination. Poly-Ubiquitin chains occur in eight basic varieties including Met1- (“linear”), K6-, K11-, K27-, K29-, K33-, K48- and K63-linked, each defined by the attachment position between one Ubiquitin and the C-terminus of an adjacent Ubiquitin. Each poly-Ubiquitin chain type adopts a distinct conformation(s) that can alter the functional properties of the newly poly-ubiquitinated target protein. Thus, when investigating the function of a poly-ubiquitinated protein, it is important to know the type and quantity of its Ubiquitin linkages.

We have partnered with IQ Proteomics to provide custom services for analysis of your ubiquitinated protein. Using the Absolute Quantitation of Ubiquitin (AQUA) technique developed in the Gygi Lab at Harvard Medical School, we can provide quantitative characterization of the different Ubiquitin linkages present in your sample.

What is AQUA?

AQUA is the Gold Standard for characterizing poly-ubiquitination. Unlike other methods, such as antibody-based Western blots, AQUA can determine both the type and relative abundance of all poly-Ubiquitin chain linkage types in protein samples. Customer-supplied samples are processed and mixed with isotopically-labeled synthetic peptide standards. The mixture is analyzed using an LC-MS/MS technique known as Parallel Reaction Monitoring (PRM). By comparing the signals from sample peptides and standards, a quantitative value is generated for each type of poly-Ubiquitin chain linkage present in the sample.

Advantages of AQUA

  • Detects and quantifies all poly-Ubiquitin chain linkage types
  • Can be scaled up or down
  • Is adaptable for detecting ubiquitination of specific target lysine(s)
 

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