Characterization of a New Marine Leucine Dehydrogenase from <i<Pseudomonas balearica</i< and Its Application for L-<i<tert</i<-Leucine Production

Leucine dehydrogenase (LeuDH) has emerged as the most promising biocatalyst for L-<i<tert</i<-leucine (L-Tle) production via asymmetric reduction in trimethylpyruvate (TMP). In this study, a new LeuDH named <i<Pb</i<LeuDH from marine <i<Pseudomonas balearica</i< w...
Ausführliche Beschreibung

Leucine dehydrogenase (LeuDH) has emerged as the most promising biocatalyst for L-<i<tert</i<-leucine (L-Tle) production via asymmetric reduction in trimethylpyruvate (TMP). In this study, a new LeuDH named <i<Pb</i<LeuDH from marine <i<Pseudomonas balearica</i< was heterologously over-expressed in <i<Escherichia coli</i<, followed by purification and characterization. <i<Pb</i<LeuDH possessed a broad substrate scope, displaying activities toward numerous L-amino acids and <i<α</i<-keto acids. Notably, compared with those reported LeuDHs, <i<Pb</i<LeuDH exhibited excellent catalytic efficiency for TMP with a <i<K</i<<sub<m</sub< value of 4.92 mM and a <i<k</i<<sub<cat</sub</<i<K</i<<sub<m</sub< value of 24.49 s<sup<−1</sup< mM<sup<−1</sup<. Subsequently, L-Tle efficient production was implemented from TMP by whole-cell biocatalysis using recombinant <i<E. coli</i< as a catalyst, which co-expressed <i<Pb</i<LeuDH and glucose dehydrogenase (GDH). Ultimately, using a fed-batch feeding strategy, 273 mM (35.8 g L<sup<−1</sup<) L-Tle was achieved with a 96.1% yield and 2.39 g L<sup<−1</sup< h<sup<−1</sup< productivity. In summary, our research provides a competitive biocatalyst for L-Tle green biosynthesis and lays a solid foundation for the realization of large-scale L-Tle industrial production. Ausführliche Beschreibung