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UbiScan analysis of TsUBE2L3 effect on the myotube ubiquitome. Open
A. Heatmap displaying proteins in which ubiquitinated peptides were found to be upregulated (blue) or downregulated (red) as measured by a fold change in relation to the empty vector C2C12 myotube cell line after 24 h expression of eGFP-HA…
Expression, purification and activity of 6His-TsUBE2L3 and 6His-ARIH2 <i>in vitro</i>. Open
A. Coommassie stain of 6His-TsUBE2L3 from un-induced (UI) and induced (I) E.coli cultures, nickel purification resin flow-through (FT1-2), wash (W) and elutions (E1-2). B and D. Streptavidin-HRP blots of in vitro parkin auto-ubiquitination…
Co-immuno-precipitation (co-IP) and yeast-2-hybrid (Y2H) analyses of TsUBE2L3. Open
A. Transgenic C2C12 myotube lines (empty vector, eGFP-HA, TsUBE2L3-HA and mouse MmUBE2L3-HA) after 24 h DOX induction were probed by co-IP with anti-HA antibody. Co-IP elutions were silver-stained and all co-IP'd proteins were analyzed by …
Identification of E2 and E3 activity in <i>T</i>. <i>spiralis</i> SP. Open
A. Streptavidin-HRP blot of in vitro ubiquitination reactions including 1) human E1 (UBE1A), E2 (UBE2L3), E3 (parkin), and Ub-biotin, 2) in the absence of the E2, 3) SP substituted for the human E2, 4) SP substituted for the human E3, 5) S…
Identification of TsUBE2L3 E3 interaction partners by co-IP and yeast-2-hybrid (Y2H) analyses. Open
Only interactions measured by Y2H as being strong are listed. Known ubiquitination substrates found by UbiScan analysis to be significantly and specifically upregulated (upreg.) or downregulated (downreg.) after expression of TsUBE2L3-HA a…
Development of expression system for TsUBE2L3 analysis in C2C12 myotubes. Open
A. Wild-type C2C12 undifferentiated myoblasts and terminally differentiated myotubes were probed by immuno-fluorescence (IFA) with anti-tubulin antibodies (Alexa-488, green) and nuclei were stained with DAPI (blue). B. Schematic of the pLV…
Verification of expression, secretion, activity and localization of TsUBE2L3. Open
A. T.sp SP, HEK 293T cell lysate and T. spiralis muscle larvae lysate were separated by SDS-PAGE and immuno-blotted with anti-HsUBE2L3 and anti-tubulin antibodies. B. Auto-ubiquitination of parkin with Ub-biotin was probed by streptavidin-…