Expanding the threonine aldolase toolbox for the asymmetric synthesis of tertiary α-amino acids
- 487 Downloads
The direct biochemical synthesis of tertiary α-amino acids with a wide range of diversity was recently reported using natural threonine aldolases LTA from Aeromonas jandei and DTA from Pseudomonas sp. Here, we describe the identification of five novel threonine aldolases which accept alanine and serine as amino acid donors. The enzymes were found by sequence database analysis using known aldolases as template. All enzymes were overexpressed in Escherichia coli and purified, and their biochemical properties were characterized. The new enantiocomplementary l- and d-threonine aldolases catalyze the asymmetric synthesis of β-hydroxy α-methyl- and α-hydroxymethyl-α-amino acids with good conversion and perfect enantioselectivity at α-carbon of the products (e.e. >99 %). The structural basis for the broad donor specificity of these threonine aldolases is analyzed based on crystal structure alignments and amino acid sequences comparison.
KeywordsThreonine aldolase Aldol reactions Tertiary amino acids Enzyme catalysis Biocatalysis
The activity leading to the present results has received funding from the European Community’s Seventh Framework Programme (FP7/2007-2013) and EFPIA companies’ in kind contribution for the Innovative Medicine Initiative under Grant Agreement No. 115360 (Chemical manufacturing methods for the 21st century pharmaceutical industries, CHEM21). We would like to thank Dr. Martina Geier for the support with genetic experiments.
This article does not contain any studies with human participants or animals performed by any of the authors. Informed consent was obtained from all individual participants included in the study.
Conflict of interest
The authors declare that they have no competing interests.
- Dietz F, Gröger H (2009) Asymmetric synthesis of all stereoisomers of α-methylthreonine using an organocatalytic steglich rearrangement reaction as a key step. Synlett 24:4208–4218Google Scholar
- Fessner WD (2011) Aldol reactions. In: Drauz K, Gröger H, May O (eds) Enzyme catalysis in organic synthesis, 3rd edn. Wiley-VCH, Weinheim, pp 857–917Google Scholar
- Grandel R, Kazmaier U (1998) Diastereoselective synthesis of β-substituted α-methylserines via alanine ester enolates. Eur J Org Chem 2:409–417Google Scholar
- Qin HM, Imai FL, Miyakawa T, Kataoka M, Okai M, Ohtsuka J, Hou F, Nagata K, Shimizu S, Tanokura M (2014) L-allo-Threonine aldolase with an H128Y/S292R mutation from Aeromonas jandaei DK-39 reveals the structural basis of changes in substrate stereoselectivity. Acta Crystallogr Sect D 70:1695–1703CrossRefGoogle Scholar