Examining ubiquitinated peptide enrichment efficiency through an epitope labeled protein

Ubiquitination is a dynamic process that is responsible for regulation of cellular responses to stimuli in a number of biological systems. Previous efforts to study this post-translational modification have focused on protein enrichment; however, recent research utilizes the presence of the di-glyci...

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Bibliographic Details
Published in:Analytical biochemistry 2016-11, Vol.512, p.114-119
Main Authors: Parker, J., Oh, Y., Moazami, Y., Pierce, J.G., Loziuk, P.L., Dean, R.A., Muddiman, D.C.
Format: Article
Language:English
Subjects:
Antibodies, Monoclonal, Murine-Derived - chemistry
Enrichment efficiency
Epitopes - chemistry
Gly-Gly enrichment
Muramidase - chemistry
Ubiquitinated Proteins - chemistry
Ubiquitination
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Summary:Ubiquitination is a dynamic process that is responsible for regulation of cellular responses to stimuli in a number of biological systems. Previous efforts to study this post-translational modification have focused on protein enrichment; however, recent research utilizes the presence of the di-glycine (Gly-Gly) remnants following trypsin digestion to immuno-enrich ubiquitinated peptides. Monoclonal antibodies developed to the cleaved ubiquitin modification epitope, (tert-butoxycarbonyl) glycylglycine (Boc-Gly-Gly-NHS)1, are used to identify the Gly-Gly signature. Here, we have successfully generated the Boc-Gly-Gly-NHS modification and showed that when conjugated to a lysine containing protein, such as lysozyme, it can be applied as a standard protein to examine ubiquitinated peptide enrichment within a complex background.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2016.08.017