Figure 5.

Metabolic activity of SAHB compounds in beta cells. (a) Panel of human SAHBs generated for islet treatment. Conserved leucine and aspartic acid residues are highlighted in yellow and serine is marked in gray. Residues altered in different SAHB compounds are marked in green and purple. N_L, norleucine. *The S5 non-natural amino acid (see Methods). (b) Effect of 3 μM SAHB on GSIS. Data are means ± s.e.m. of five independent experiments. Asterisks compare release at 5.5 mM versus 12.5 or 25 mM glucose within each group of islets, **P < 0.01, ***P < 0.001. ^‡P < 0.05 or P < 0.001, Bad ^+/+ versus Bad ^−/− islets treated with vehicle. ^†P < 0.05 or P < 0.01, Bad ^−/− islets treated with vehicle versus BAD SAHB_A. ^¶P < 0.05 or P < 0.01, Bad ^−/− islets treated with BAD SAHB_A versus phosphomimetic SAHBs (SAHB_A(S→pS) or SAHB_A(S→D)). Insulin content per islet was 114.61 ± 5.18 and 105.54 ± 4.63 ng, Bad ^+/+ and Bad ^−/−, respectively. (c) Effect of 1 μM SAHB compounds on glucose-induced changes in ΔΨ_m. (d) Binding affinities of SAHBs to recombinant BCL-X_L ΔC protein. (e) Effect of 3 μM SAHBs on glucokinase activity in INS-1 cells. *P < 0.05 and ***P < 0.001. (f) Panel of derivatized mouse SAHBs containing a photoactivatable benzophenone moiety. (g) Identification of glucokinase as a direct BAD BH3 target. Formation of a covalent complex between SAHB_A(S→pS) and glucokinase upon UV photoactivation. (h) Competition of SAHB compounds with full-length IVTT BAD for binding to IVTT glucokinase. IP, immunoprecipitation.