Abstract
A novel D-lyxose isomerase has been identified within the genome of a hyperthermophilic archaeon belonging to the Thermofilum species. The enzyme has been cloned and over-expressed in Escherichia coli and biochemically characterised. This enzyme differs from other enzymes of this class in that it is highly specific for the substrate D-lyxose, showing less than 2% activity towards mannose and other substrates reported for lyxose isomerases. This is the most thermoactive and thermostable lyxose isomerase reported to date, showing activity above 95°C and retaining 60% of its activity after 60 min incubation at 80°C. This lyxose isomerase is stable in the presence of 50% (v/v) of solvents ethanol, methanol, acetonitrile and DMSO. The crystal structure of the enzyme has been resolved to 1.4–1.7 A. resolution in the ligand-free form and in complexes with both of the slowly reacting sugar substrates mannose and fructose. This thermophilic lyxose isomerase is stabilised by a disulfide bond between the two monomers of the dimeric enzyme and increased hydrophobicity at the dimer interface. These overall properties of high substrate specificity, thermostability and solvent tolerance make this lyxose isomerase enzyme a good candidate for potential industrial applications.
Highlights
In the last 50 years, the incidence of chronic “lifestyle” diseases such as diabetes, obesity, hyperlipidemia, and hypertension has increased rapidly throughout the world
To identify thermophilic LIs BLAST (Altschul et al, 1990) searches in genomes from hyperthermophilic archaea and bacteria, and metagenomes collected from thermophilic habitats, were performed using the sequences of biochemically and structurally characterized D-LIs as templates
A putative thermophilic LI was identified within the metagenomic sequences isolated from deep-sea hydrothermal vents and assigned to the Thermofilum species ex4484_79 (Locus tag: B6U94_07925)
Summary
In the last 50 years, the incidence of chronic “lifestyle” diseases such as diabetes, obesity, hyperlipidemia, and hypertension has increased rapidly throughout the world. These diseases are generally caused by the over intake of high sugar and high-fat foods. Rare sugars can be important building blocks for new drugs (Kwon et al, 2010). Chemical synthesis of rare sugars requires strict reaction conditions, complicated purification steps, produces chemical waste, and has production safety issues. The application of a biocatalytic route for sugar production is more sustainable it requires the application of specific enzymes. The stability of the required enzymes can be a limitation for their commercial
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