N-succinylamino acid racemases: Enzymatic properties and biotechnological applications

Abstract

amino acid racemase; racemase; amino acid; enzymatic cascade Abbreviations: N-succinylamino acid racemase (NSAR); o-succinylbenzoate synthase (OSBS); N-succinylamino acid racemase/o-succinylbenzoate synthase subfamily (NSAR/OSBS); N-acetyl-amino or N-acyl-amino acid racemase (NAAR); Nsubstituted-amino acid (NxA); N-substituted-amino acid racemase (NxAR); Kinetic resolution (KR); Dynamic kinetic resolution (DKR); N-acetyl-(NA); N-acetyl-amino acid (NAA); N-succinyl-(NS); N-succinyl-amino acid (NSA); N-carbamoyl-(NC); Ncarbamoyl-amino acid (NCAA); N-chloroacetyl-(NCh); N-chloroacetyl-amino acid (NChAA); N-formyl-(NF); N-formyl-amino acid (NFAA); N-butyril (NBt); N-butyril amino acid (NBtA); N-propionyl (NPr); N-propionyl amino acid (NPrA); N-benzoyl (NBzA); N-benzoyl amino acid (NBzA); (1R,6R)-2-succinyl-6-hydroxy-2,4cyclohexadiene-1-carboxylate (SHCHC); Amycolatopsis sp. TS-1-60 NSAR (AmyNSAR); Geobacillus kaustophilus NSAR (GkaNSAR); Geobacillus stearothermophilus CECT 49 NSAR (GstNSAR); Streptomyces atratus Y-53 NSAR (SatNSAR); Sebekia benihana NSAR (SebeNSAR); Chloroflexus aurantiacus NSAR (CauNSAR); Thermus thermophilus NSAR (TteNSAR); Deinococcus radiodurans NSAR (DraNSAR); Exiguobacterium sp. AT1b OSBS (ExiOSBS); Alicyclobacillus acidocaldarius OSBS (AacOSBS); Enterococcus faecalis V583 NSAR (EfaNSAR); Listeria innocua Clip11262 NSAR (LinNSAR); Lysinibacillus varians GY32 NSAR (LvaNSAR); Roseiflexus castenholzii HLO8 NSAR (RcaNSAR); Amycolatopsis mediterranei S699 NSAR (AmedNSAR).

Overview on Multienzymatic Cascades for the Production of Non-canonical α-Amino Acids

“…Whole cell systems containing NSAR and L-succinylase from Geobacillus kaustophilus were successfully applied for the synthesis of L-4-bromophenylalanine, L-3fluorophenylalanine, L-2-naphthylalanine, L-2-indanylglycine, and L-6-heptenylglycine (conversion over 98%, >99.9% e.e.) ( Figure 2D, Masutoshi et al, 2016). An NSAR/D-succinylase system has also been reported (Sumida et al, 2016(Sumida et al, , 2018.…”

“…Different combinations of purified and immobilized D-hydantoinase (DHyd) and hydantoin racemase (HR) from Agrobacterium species, N-succinyl-amino acid racemase (NSAR) from Geobacillus kaustophilus and L-carbamoylase (LCar) from Geobacillus stearothermopillus allowed the production of different enantiopure NcAAs; L-Met and L-Val were also efficiently produced with this system (Rodríguez-Alonso et al, 2015, 2016, 2017. (Martínez-Rodríguez et al, 2020), and hydantoin racemases (Martínez-Rodríguez et al, 2004). This process has also been expanded for the production of enantio-enriched β-AAs (Martínez-Gómez et al, 2012;Rudat et al, 2012).…”

“…R: formyl, NAAR/carbamoylase tandem. R: succinyl, NSAR/succinylase tandem (Martínez-Rodríguez et al, 2020). (B) L-homophenylalanine production by NSAR/L-acylase tandem.…”

“…(C) D-allylglycine production by NSAR/D-acylase tandem, consisting of pure D-acylase (D-Acyl, Chirotech Technology Ltd.) and an engineered N-succinyl-amino acid racemase from Amycolatopsis sp (NSAR, G291D/F323Y mutant; Baxter et al, 2012). (D) Production of enantiopure L-α-AA using an NSAR/L-succinylase system, using enzymes from Geobacillus kaustophilus overexpressed in a whole cell system (Masutoshi et al, 2016). (E) Production of enantio-enriched D-Phe and D-Trp using a NSAR/D-succinylase system.…”

“…The “ D -system” appears contained in the dashed line, whereas the “ L -system” is remarked by a full line. Inclusion of an NSAR enzyme highly improves the “ L -system.” HR*: hydantoin racemase (or chemical racemization when favored); DHyd, D -activity of hydantoinase; DCar, D -carbamoylase; LHyd, L -activity of hydantoinase; LCar, L -carbamoylase; NSAR, N -carbamoyl-racemase promiscuous activity of N -succinyl-racemase ( Martínez-Rodríguez et al, 2010b , 2020 ). D -AA, D -α-amino acid.…”