The stereoselectively liganded targets of 3-amino stereoprobes span a range of protein classes including nucleotide exchange factors (DENND2D), methyltransferases (TRMT61A) and other proteins responsible for RNA modification (MTO1) and transport (CLUH), scaffolding subunits of metabolic enzymes (UQCRC1), glutathione S-transferases (GSTO1), and kinases (IKBKB, STK39) (see SI Figure for full list). In Figure -G, we highlight the stereoselective enrichment of UQCRC1 by 36B and MTO1 and IKBKB by 36D, and blockade of enrichment by 37B and 37D, respectively. None of these proteins showed strong evidence of stereoselective enrichment by the 3-aryl stereoprobes. In summary, in comparison to previously described 3-aryl-azetidine acrylamides, the 3-amino-azetidine acrylamides displayed lower overall proteomic reactivity that nonetheless included discrete and unique stereoselective liganding events, highlighting the value of gaining synthetic access to structurally diverse azetidine stereoprobes. Enantioselectivity is defined as the ratio of enrichment for one stereoisomer versus its enantiomer, and diastereoselectivity as the ratio of enrichment of one stereoisomer versus the average of its two diastereomers. For the quadrant plots, a protein is shown in color if both the enantioselectivity and diastereoselectivity of enrichment (log2) are greater than 1. (C) Pie chart showing the total number of proteins stereoselectively liganded by each stereoisomer of 3-amino-azetidine acrylamide stereoprobes. A protein is defined as being stereoselectively enriched if the enantioselectivity is greater than 3-fold; a protein was designated as being "liganded" if stereoselectively enriched and if pretreatment by the corresponding competitor probe resulted in >50% blockade of this enantioselective enrichment as compared to pretreatment with DMSO. (D) Bar graph comparing the total number of proteins stereoselectively enriched (lighter tan) and liganded (darker tan) by 3-amino stereoprobes vs 3-aryl stereoprobes in Ramos cells (treated with 5 µM of alkynylated 3-aryl stereoprobes and 10 µM of alkynylated 3-amino stereoprobes; pretreatment with 20 µM of either 3-aryl-or 3-amino competitor stereoprobes). A stereoselectively enriched protein was considered "shared" by the 3-amino and 3-aryl stereoprobes if it was enriched with >3-fold enantioselectivity by one stereoprobe chemotype and >2-fold enantioselectivity by the other stereoprobe chemotype. A liganded protein was considered "shared" if it was stereoselectively enriched and competed by >50% by one stereoprobe chemotype and >30% by the other stereoprobe chemotype. All other stereoselectively enriched or liganded targets of the 3-amino and 3-aryl stereoprobes were considered chemotype-selective. (E, F, G) Bar graphs showing protein-directed ABPP data for UQCRC1, MTO1, and IKBKB each of which was stereoselectively liganded by 3-amino stereoprobes, but not 3-aryl stereoprobes.