Extended Data Fig. 2: Biotinylated macrocycles pull down proteasome subunits in cell lysates.
From: Targeted degradation via direct 26S proteasome recruitment
a, Affinity purification of L- and D- versions of MC1biotin incubated with HEK 293 lysates run on SDS-PAGE and stained for total protein elution. b, Western blot showing MC1biotin, but not the inactive D enantiomer or DMSO, co-purifies with 19S subunits PSMD1, PSMD4, as well as 20S subunit PSMA1. c, The distribution of normalized log2 protein intensities across three biological replicates from each condition shown in (a) and (b) show similar abundance across all samples. Box plots represent data distribution (minimum, first quantile, median, third quantile, and maximum) with all proteins. The lower whisker is the smallest normalized log2 intensities or equal to lower bound of box-1.5*IQR (Interquantile range=75% quantile-25% quantile). The upper whisker is the largest normalized log2 intensities or equal to upper bound of box+1.5*IQR. Outliers are displayed as dots in each sample. d, Sample correlation heatmap showing correlation between each biological replicate, represented by Pearson coefficient. e-g, Volcano plots showing enrichment for L- vs D- affinity purification (panel e, included from Fig. 1d for comparison), L- vs -DMSO (f), and DMSO vs D- affinity purification (g), showing clear enrichment of proteasome subunits for the L-enantiomer of MC1. Data in e-g represent three biological replicates per condition (L-CIDE, D-CIDE and DMSO control). Horizontal dotted line shows p-value=0.05. Vertical dotted lines show log2(fold change)= -1 or 1. For details of data and statistical analysis see methods.
