2024-03-29T02:33:13Zhttp://digital.csic.es/dspace-oai/requestoai:digital.csic.es:10261/161832020-07-03T11:27:23Zcom_10261_5063com_10261_5col_10261_5066
Recombinant production of serine hydroxymethyl transferase from Streptococcus thermophilus and its preliminary evaluation as a biocatalyst
Vidal, Luis
Calveras Ibáñez, Jordi
Clapés Saborit, Pere
Ferrer, Pau
Caminal, Glòria
Serine hydroxymethyl transferase
Streptococcus thermophilus
glyA gene
9 pages, 3 figures, 3 tables.-- PMID: 15726349 [PubMed].-- Printed version published Sep 2005.
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 K m 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.
Financial support from CICYT (project PPQ 2002-04625-C02-01).
Peer reviewed
2009-08-24T08:55:02Z
2009-08-24T08:55:02Z
2005-02-23
artículo
http://purl.org/coar/resource_type/c_6501
Applied Microbiology and Biotechnology 68(4): 489-497 (2005)
0175-7598
http://hdl.handle.net/10261/16183
10.1007/s00253-005-1934-1
1432-0614
en
http://dx.doi.org/10.1007/s00253-005-1934-1
none
22195 bytes
application/pdf
Springer