TUBE-Based Mass Spectrometry
Workflow schematic of TUBE proteomics. Treated cells are enriched using TUBEs (TUBE1 and K48). The enriched polyubiquitylated proteins are pixelated by SDS-PAGE and digested with trypsin. Individual ubiquitylated proteins are identified by the presence of at least 2 peptides in at least 2 of 3 replicates. Ub linkage types are identified by K-ε-GG peptides corresponding to covalent Ub-Ub modification at a particular lysine.
TUBE-Based Mass Spectrometry Proteomics
Tandem Ubiquitin Binding Entities (TUBEs) are powerful reagents for enrichment of polyubiquitylated proteins. At LifeSensors, we have used this remarkable technology of TUBEs with our mass spectrometry expertise to provide the customer with a quick and easy way to perform both qualitative and quantitative proteomics. Upon completion of the analysis, you will receive a list of peptides and corresponding proteins that have been identified. We will categorize the ubiquitylated proteins and display the results on several plots to make your analysis easier. This information will help you publish your results or plan your next assay.
Tandem Ubiquitin Binding Entities (TUBEs) are powerful reagents for enrichment of polyubiquitylated proteins. At LifeSensors, we have used this remarkable technology of TUBEs with our mass spectrometry expertise to provide the customer with a quick and easy way to perform both qualitative and quantitative proteomics. Upon completion of the analysis, you will receive a list of peptides and corresponding proteins that have been identified. We will categorize the ubiquitylated proteins and display the results on several plots to make your analysis easier. This information will help you publish your results or plan your next assay.
We are here to help you from the beginning, and are happy to sit down and help you design the best experiment to address your specific questions. We have competitively priced our analysis and offer custom services with the level of analysis that matches your needs and budget. We strive to deliver you accurate and professional reports quickly. From submission of samples reports are typically delivered in 2-4 weeks depending on the complexity of project. Please contact us for a more accurate timeline for your specific experiment.
TUBE-based enrichment of polyubiquitylated proteins has proven key for the progress of ubiquitin proteomics. LifeSensors has developed the proprietary technology to identify cellular proteins that are ubiquitylated using TUBE-based proteomics. A brief overview of this technology is depicted in the following figure:
See Sample Report
Chain Selectivity
EXPERIMENTAL DATA
In-house data
Below are examples of data generated using TUBE- based mass spec technology with pan- selective TUBEs. Here the ubiquitinated proteins have been differentiated from whole cell lysate. This methodology is sensitive enough to detect the number of ubiquitin sites on a given protein. The number of ubiquitinated proteins detected was assigned to the number of ubiquitin sites that they carry. In this case over half of the ubiquitnated proteins detected were monoubiquitinated. When studying the effects of a drug or genetic knockout global shifts in the ubiquitome is a good place to start your analysis.
PUBLISHED DATA
In 2016, Mata-Cantero et al.2 used TUBE-based mass-spectrometry to identify major components of the ubiquitin proteome of both Plasmodium falciparum and its host during different life stages.
Identification of ubiquitylated proteins using TUBEs-LC-MS/MS method. Intraerythrocytic Developmental Cycle of P. falciparum is shown. Synchronized P. falciparum iRBC at 40% parasitaemia from rings, trophozoites and schizonts stages were collected and frozen. TUBE enriched proteins from iRBC at different stages and uRBC were captured using TUBEs or GST (control) previously crosslinked with DMP to agarose beads. After exhaustive washes, proteins captured were eluted, cleaned by precipitation and resolved by electrophoresis (PAGE). Bands with proteins were analyzed by LC-MS/MS.