The incorporation of organocatalysts into protein scaffolds holds the promise of overcoming some of the limitations of this powerful catalytic approach. Previously, we showed that incorporation of the non-canonical amino acid para-aminophenylalanine into the non-enzymatic protein scaffold LmrR forms a proficient and enantioselective artificial enzyme (LmrR_pAF) for the Friedel-Crafts alkylation of indoles with enals. The unnatural aniline side-chain is directly involved in catalysis, operating via a well-known organocatalytic iminium-based mechanism. In this study, we show that LmrR_pAF can enantioselectively form tertiary carbon centres not only during C-C bond formation, but also by enantioselective protonation, delivering a proton to one face of a prochiral enamine intermediate. The importance of various side-chains in the pocket of LmrR is distinct from the Friedel-Crafts reaction without enantioselective protonation, and two particularly important residues were probed by exhaustive mutagenesis.
Keywords: Artificial Enzymes; Enantioselective Protonation; Friedel-Crafts Alkylations; Iminium Catalysis; Non-canonical Amino Acids.
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