Hyperthermophilic aldolases as biocatalyst for C–C bond formation: rhamnulose 1-phosphate aldolase from Thermotoga maritima

• Published in: Applied microbiology and biotechnology Vol. 99; no. 7; pp. 3057 - 3068
• Main Authors: Oroz-Guinea, Isabel, Sánchez-Moreno, Israel, Mena, Montaña, García-Junceda, Eduardo
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01.04.2015; Springer; Springer Nature B.V
• Subjects: Aldehyde-Lyases - chemistry; Aldehyde-Lyases - genetics; Aldehyde-Lyases - metabolism; Aldolases; Amino Acid Sequence; Analysis; Biocatalysts; Biomedical and Life Sciences; Biotechnologically Relevant Enzymes and Proteins; Biotechnology; Catalysis; Cloning, Molecular; E coli; Enzyme Stability; Enzymes; Escherichia coli; Escherichia coli - genetics; Experiments; Half-Life; Hydrogen-Ion Concentration; Influence; Kinases; Kinetics; Life Sciences; Microbial Genetics and Genomics; Microbiology; Microorganisms; Molecular Sequence Data; Physiological aspects; Recombinant Proteins - genetics; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Studies; Temperature; Thermotoga maritima; Thermotoga maritima - enzymology; Thermotoga maritima - genetics; Zinc compounds; Spain; Thermostability; Aldolases; Biocatalysis; Hyperthermophilic enzymes; Thermozyme; Enzyme catalysis
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-014-6123-7

The TM1072 gene from Thermotoga maritima codifies for a putative form of a rhamnulose-1-phosphate aldolase (Rha-1PA Tm). To investigate this enzyme further, its gene was cloned and expressed in Escherichia coli . The purified enzyme was activated by Co 2+ as a divalent metal ion cofactor, instead of Zn 2+ as its E. coli homologue, and exhibited a maximum of activity at 95 °C. Furthermore, the enzyme displayed a high stability against extreme reaction conditions, retaining 90 % of its activity in the presence of 40 % of acetonitrile and showing a half-life greater than 3 h at 115 °C. The kinetic parameters at room temperature (R/T) were also studied; the K M was calculated to be 3.6 ± 0.33 mM, while k cat / K M was found to be 0.7 × 10 3  s −1  M −1 . Given these characteristics, Rha-1PA Tm is an attractive enzyme for use as a biocatalyst for industrial applications, offering intriguing possibilities for practical biocatalysis.