Applied Microbiology and Biotechnology

, Volume 68, Issue 4, pp 489–497

Recombinant production of serine hydroxymethyl transferase from Streptococcus thermophilus and its preliminary evaluation as a biocatalyst

Authors

  • L. Vidal
    • Unitat de Biocatalisi Aplicada Associada al IIQAB (CSIC-UAB), Departament d’Enginyeria Química, Escola Tècnica Superior d’EnginyeriaUniversitat Autònoma de Barcelona
  • J. Calveras
    • Institute for Chemistry and Environmental Research—CSIC
  • P. Clapés
    • Institute for Chemistry and Environmental Research—CSIC
  • P. Ferrer
    • Unitat de Biocatalisi Aplicada Associada al IIQAB (CSIC-UAB), Departament d’Enginyeria Química, Escola Tècnica Superior d’EnginyeriaUniversitat Autònoma de Barcelona
    • Unitat de Biocatalisi Aplicada Associada al IIQAB (CSIC-UAB), Departament d’Enginyeria Química, Escola Tècnica Superior d’EnginyeriaUniversitat Autònoma de Barcelona
Biotechnologically Relevant Enzymes and Proteins

DOI: 10.1007/s00253-005-1934-1

Cite this article as:
Vidal, L., Calveras, J., Clapés, P. et al. Appl Microbiol Biotechnol (2005) 68: 489. doi:10.1007/s00253-005-1934-1

Abstract

The glyA gene encoding a serine hydroxymethyl transferase (SHMT) with threonine aldolase activity was isolated from Streptococcus thermophilus YKA-184 chromosomal DNA. This aldolase is a pyridoxal 5′-phosphate-dependent enzyme that stereospecifically catalyzes the interconversion of l-threonine to glycine and acetaldehyde. The enzyme was overexpressed in Escherichia coli M15 as a recombinant protein of 45 kDa with a His6-tag at its N-terminus. The recombinant enzyme was purified to homogeneity by a single chromatographic step using Ni-nitrilotriacetic acid affinity, obtaining a high activity-recovery yield (83%). Lyophilized and precipitated enzymes were stable at least for 10 weeks when stored at −20°C and 4°C. It was observed that the Km for l-allo-threonine was 38-fold higher than that for l-threonine, suggesting this enzyme can be classified as a specific l-allo-threonine aldolase. The optimum pH range of threonine aldolase activity for the recombinant SHMT was pH 6–7. When tested for aldol addition reactions with non-natural aldehydes, such as benzyloxyacetaldehyde and (R)-N-Cbz-alaninal, two possible β-hydroxy-α-amino acid diastereoisomers were produced, but with moderate stereospecificity. The enzyme showed potential as a biocatalyst for the stereoselective synthesis of β-hydroxy-α-amino acids.

Copyright information

© Springer-Verlag 2005