Threonine aldolases: perspectives in engineering and screening the enzymes with enhanced substrate and stereo specificities.

Kateryna Fesko

doi:10.1007/s00253-015-7218-5

Abstract

Threonine aldolases have emerged as a powerful tool for asymmetric carbon-carbon bond formation. These enzymes catalyse the unnatural aldol condensation of different aldehydes and glycine to produce highly valuable β-hydroxy-α-amino acids with complete stereocontrol at the α-carbon and moderate specificity at the β-carbon. A range of microbial threonine aldolases has been recently recombinantly produced by several groups and their biochemical properties were characterized. Numerous studies have been conducted to improve the reaction protocols to enable higher conversions and investigate the substrate scope of enzymes. However, the application of threonine aldolases in organic synthesis is still limited due to often moderate yields and low diastereoselectivities obtained in the aldol reaction. This review briefly summarizes the screening techniques recently applied to discover novel threonine aldolases as well as enzyme engineering and mutagenesis studies which were accomplished to improve the catalytic activity and substrate specificity. Additionally, the results from new investigations on threonine aldolases including crystal structure determinations and structural-functional characterization are reviewed.

Highlights

Threonine serves as the sole source of carbon and nitrogen for the growth of a wide variety of organisms
Some enzymes participate in the biosynthesis of threonine from aspartic acid via homoserine, whereas others are responsible for the degradation of threonine (Scheme 1)
This review briefly summarizes the screening and selection methods used in search of new threonine aldolases as well as new structural-functional knowledge which might be helpful for the design of new activities

Summary

Introduction

Threonine serves as the sole source of carbon and nitrogen for the growth of a wide variety of organisms. Besides TAs, another PLPdependent enzyme, the serine hydroxymethyltransferase (SHMT), catalyses the retro-aldol cleavage of L-βhydroxy-α-amino acids similar to LTAs in an alternative route (Vidal et al 2005; Gutierrez et al 2008). The currently described protocols for the enzymatic synthesis of β-hydroxy-α-amino acids suffer from a limited substrate range allowing only glycine as the donor, moderate yields and low stereoselectivities at the β-carbon (diastereoselectivity, d.e.) of the product. The limitation of rigid donor specificity was recently overcome by isolation of natural TAs with broad donor specificity LTA from Aeromonas jandaei and DTA from Pseudomonas sp., which accept alanine and serine as donors; L- and D-α-quaternary-α-amino acids were produced (Fesko et al 2010).

Summary and outlook

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