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. Author manuscript; available in PMC: 2016 Feb 16.
Published in final edited form as: J Am Chem Soc. 2015 Aug 25;137(35):11303–11311. doi: 10.1021/jacs.5b04366

Figure 2.

Figure 2

Structure-based design of a fluorogenic probe for folded and functional RA. (a) Schematic of a push-pull environmentally-sensitive fluorophore. EWG = electron-withdrawing group. EDG = electron-donating group. (b) Structure of the retroaldol substrate S1 utilized by the de novo designed RA enzyme.29 RA catalyzes a retroaldol reaction using the pKa-perturbed lysine-210 ε-amine side chain that forms a Schiff base with S1. (c) P1 is a push-pull environmentally-sensitive fluorophore featuring a reactive vinyl ketone (in red) that also serves as an electron-withdrawing group. The dimethyl amino group (in green) serves as an electron-donating group. P1 covalently modifies the pKa-perturbed lysine-210 residue of RA through 1,4-conjugate addition, rendering the RA-P1 covalent conjugate fluorescent.