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

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

Automated summary

In this study, a new LeuDH named PbLeuDH was heterologously over-expressed in Escherichia coli, followed by purification and characterization. PbLeuDH exhibited excellent catalytic efficiency for TMP and was used for high-yield L-Tle production through whole-cell biocatalysis. This research provides a competitive biocatalyst for green biosynthesis of L-Tle.