Fs hydrodex β 6tbdm

All chemicals were purchased from Sigma-Aldrich (Steinheim, Germany). Arylmalonic acids 3b, 3c and 3d were kindly provided by Chiracon GmbH (Teltow, Germany). Fluorescence and absorption were measured with a FLUOstar Omega Fluorimeter (BMG Labtech GmbH Ortenberg, Germany) DNA samples were sequenced by the DNA sequencing service of the department of biochemistry at Ruhr-University Bochum. Kits for ligation, DNA isolation and restriction were purchased at Fermentas (St. Leon-Rot, Germany). Enzyme affinity purification was done using a 1 mL His Pure Ni-NTA column (Pierce Biotechnology Rockford, USA). Chiral GC analyses were carried out using the chiral column FS-Hydrodex-β-6TBDM [heptakis-(2,3-di-O-methyl-6-O-t-butyldimethylsilyl)-β-cyclodextrin] (Macherey Nagel, Germany) on a GC-FID-2010 (Shimadzu, Japan). Retention times for 2b (160°C) were 31.5 min (S) and 32.3 min (R) for 2c (170°C) 12.9 min (S) and 14.4 min (R). The elution order was identified using commercial (S)-naproxen (TCI Chemicals, Japan). Chiral HPLC analyses were carried out using the column CHIRALCEL OD-H (Daicel, Osaka, Japan) on a L-7110 HPLC device (Hitachi High Tech, Tokyo, Japan) using n-hexane/i-propanol/trifluor acetic acid (98:2:0.5). With a flow rate of 0.5 mL, the two enantiomers of phenyl methyl malonic acid ethyl ester 2a eluted with 55 min (S) and 65 min (R).

Substrates 1 were synthesized as described in the Supporting Information. All other chemicals used in this study were purchased from commercial suppliers. A Bruker (Rheinstetten, Germany) DPX‐200 NMR device was used for ¹H and ¹³C NMR measurements. High performance liquid chromatography (HPLC) analyses were performed with an AZURA HPLC System (Knauer, Berlin, Germany) with a NUCLEODUR C18 Pyramid column (5 μm; 4.6×250 mm; Macherey–Nagel, Düren, Germany). The enantiomeric excess of intermediates 2 and products 3 was determined by chiral gas chromatography with a flame ionization detector (GC‐FID) by using a Shimazu GC Plus 2010 device (Shimazu, Kyoto, Japan) with a chiral column FS‐Hydrodex‐β‐6TBDM (Macherey–Nagel, Düren, Germany).
The codon‐optimized genes of B. bronchiseptica AMD‐WT (UniProtKB/Swiss‐Prot: Q05 115, PDB: 3DG9) and its variants V43I/A125P/V156L/M159L (AMD‐IPLL), G74C/M159L/C188G (AMD‐CLG), and V43I/G74C/A125P/V156L/M159L/C188G (AMD‐CLGIPL) were cloned into a pET28a vector with treatment of restriction enzymes NdeI, and XhoI. E. coli BL21(DE3) competent cells (Agilent Technologies, Santa Clara, CA, USA) were used for protein expression.22