Profiling lysine ubiquitination by selective enrichment of ubiquitin remnant-containing peptides.

Protein ubiquitination plays critical roles in many biological processes. However, functional studies of protein ubiquitination in eukaryotic cells are limited by the ability to identify protein ubiquitination sites. Unbiased high-throughput screening methods are necessary to discover novel ubiquitination sites that play important roles in cellular regulation. Here, we describe an immunopurification approach that enriches ubiquitin remnant-containing peptides to facilitate downstream mass spectrometry (MS) identification of lysine ubiquitination sites. This approach can be utilized to identify ubiquitination sites from proteins in a complex mixture.