The identification of a robust leucine dehydrogenase from a directed soil metagenome for efficient synthesis of L-2-aminobutyric acid.
Biotechnol J
; 18(8): e2200590, 2023 Aug.
Article
in En
| MEDLINE
| ID: mdl-37149736
ABSTRACT
L-2-aminobutyric acid (L-2-ABA) is a chiral precursor for the synthesis of anti-epileptic drug levetiracetam and anti-tuberculosis drug ethambutol. Asymmetric synthesis of L-2-ABA by leucine dehydrogenases has been widely developed. However, the limitations of natural enzymes, such as poor stability, low catalytic efficiency, and inhibition of high-concentration substrates, limit large-scale applications. Herein, by directed screening of a metagenomic library from unnatural amino acid-enriched environments, a robust leucine dehydrogenase, TvLeuDH, was identified, which exhibited high substrate tolerance and excellent enzymatic activity towards 2-oxobutyric acid. In addition, TvLeuDH has strong affinity for NADH. Subsequently, a three-enzyme co-expression system containing L-threonine deaminase, TvLeuDH, and glucose dehydrogenase was established. By optimizing reaction conditions, 1.5 M L-threonine could be converted to L-2-ABA with a 99% molar conversion rate and a space-time yield of 51.5 g·L-1 ·h-1 . In this process, no external coenzyme was added. The robustness of TvLeuDH allowed the reaction to be performed without the addition of extra salt as the buffer, demonstrating the simplest reaction system currently reported. These unique properties for the efficient and environmentally friendly production of chiral amino acids make TvLeuDH a particularly promising candidate for industrial applications, which reveals the great potential of directed metagenomics for industrial biotechnology.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Metagenome
/
Aminobutyrates
Type of study:
Diagnostic_studies
Language:
En
Journal:
Biotechnol J
Journal subject:
BIOTECNOLOGIA
Year:
2023
Document type:
Article
Affiliation country:
China