Journal
MOLECULAR CATALYSIS
Volume 525, Issue -, Pages -Publisher
ELSEVIER
DOI: 10.1016/j.mcat.2022.112355
Keywords
L-Threonine Aldolase; C-beta-stereoselectivity; Prereaction State; Per-residue Energy Decomposition; Esterification
Categories
Funding
- National Natural Science Foundation of China [22108039]
- Open Test Fund for Precious Instruments of Fuzhou University [2022T017]
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By rational design, researchers successfully screened a mutant LTA with enhanced C-beta-stereoselectivity, improving the efficiency of synthesizing (2S, 3R)-1b. Furthermore, esterification was used to successfully synthesize high-purity (2S, 3R)-1d, providing an effective method for green synthesis.
L-threo-4-methylsulfonylphenylserine ethyl ester [(2S, 3R)-1d], a key building block of florfenicol which synthesized by esterification using ethanol and L-threo-4-methylsulfonylphenylserine [(2S, 3R)-1b] as substrates. L-threonine aldolase (LTA) is a promising biocatalyst for producing (2S, 3R)-1b through a one-step process taking 4-Methylsulphonyl benzaldehyde (1a) and glycine as substrates under the mild condition. However, the moderate C-beta-stereoselective blocked the industrial application of LTA. To address this issue, rational design by combination with prereaction state molecular dynamics (MD) simulation and per-residue energy decomposition algorithm are employed to engineer LTA for enhancing C-beta-stereoselective. As a result, a triple mutant N16A/E98S/Y314R (Mu3) is screened out to produce (2S, 3R)-1b with 93.7% de and 90.2% conversion under the 300 mM 1a substrate loading. Furthermore, esterification is applied to synthesize (2S, 3R)-1d with over 99% chiral purity and 99% product purity. The success of this study provides new insights for the rational design of LTA with improved C-beta-stereoselectivity and proves a chemo-enzymatic route for green synthesis of (2S, 3R)-1d.
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