In an initial screen of commercial wild-type transaminases using L-alanine (L-Ala) as an amine donor, PRO-TRANS(248) provided 3 in >99% enantioselectivity, albeit with low conversion (Table , entry 1). Neither increasing enzyme loading (entry 2) nor L-Ala stoichiometry (entries 3 and 4) resulted in conversions greater than 50%, suggesting that the amine transfer equilibrium favors L-Ala and ketoacid 2 (Keq ~0.046, vide infra). Encouraged by the promiscuity of PRO-TRANS(248) toward other amino acid donors, we hypothesized that lysine could induce a more favorable equilibrium via spontaneous cyclization of the 6-amino-2-oxohexanoic acid byproduct. Indeed, use of only 1.3 and 4.0 equiv. L-lysine (L-Lys) achieved 81% and 99% conversion, respectively, over 24 h (entries 5 and 6). Increasing enzyme loading to 75 wt% and increasing reaction time by an additional 24 h led to full conversion (entry 7). The preparative value of this reaction was demonstrated on 100 g scale using 32 wt% PRO-TRANS(248) to afford 3 as a single isomer in 80% isolated yield after a simple precipitation from the reaction mixture (entry 8, see Supporting Information for details). Substrate Scope. With optimized conditions identified for the DKR of 2 to 3 (82% AY, >20:1 dr), we evaluated the -ketoacid substrate scope of PRO-TRANS(248) to provide access to -naphthyl alanine (3a) and a variety of -branched ncAAs on 0.5-2.0 mmol scale (Figure ). The enzyme is tolerant of substitution on the -naphthyl ring (3b -3d), affording (S,S)-ncAAs with >20:1 dr and moderate to excellent yields (see Supporting Information for selectivity analysis); moving the ncAA branch point to another naphthyl position (2-naphthyl, 3e) led to a measurably lower dr (9:1), despite good yield. High diastereoselectivities (>20:1) were restored for substrates with substituted -phenyl rings (3f, 3g), and -heteroaryl groups (3h-3l). Impressively, PRO-TRANS(248) is tolerant of larger -alkyl functionality, including n-propyl (3m), isopropyl (3n), Reactions were performed at 0.5-2.0 mmol scale at 20 g/L substrate loading with 1.5 equiv. L-Lys and 50 wt% Pro-TRANS(248) in 100 mM borate buffer at 37 °C for 48 h. In parentheses: yields refer to the isolated product after purification by reverse phase column chromatography (see Supporting Information for details). The single crystal of 3 was grown as a dihydrate; water was omitted from ORTEP for clarity. The single crystal of 3p was grown as an HCl salt; the chloride was omitted from ORTEP for clarity. a Reaction was conducted at pH 8.2 to avoid ester hydrolysis. b Product was isolated as the Fmoc derivative for purification purposes.