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Abstract A novel l -rhamnose isomerase was identified and cloned from an extreme-temperature aquatic habitat metagenome. The deduced amino acid sequence homology suggested the possible source of this metagenomic sequence to be Chloroflexus islandicus . The gene expression was performed in a heterologous host, Escherichia coli , and the recombinant protein l -rhamnose isomerase (L-RI M ) was extracted and purified. The catalytic function of L-RI M was characterized for d -allulose to d -allose bioconversion. d -Allose is a sweet, rare sugar molecule with anti-tumour, anti-hypertensive, cryoprotective, and antioxidative properties. The characterization experiments showed L-RI M to be a Co ++ - or Mn ++ -dependent metalloenzyme. L-RI M was remarkably active (~ 80%) in a broad spectrum of pH (6.0 to 9.0) and temperature (70 to 80 °C) ranges. Optimal L-RI M activity with d -allulose as the substrate occurred at pH 7.0 and 75 °C. The enzyme was found to be excessively heat stable, displaying a half-life of about 12 days and 5 days at 65 °C and 70 °C, respectively. L -RI M catalysis conducted at slightly acidic pH of 6.0 and 70 °C achieved biosynthesis of about 30 g L −1 from 100 g L −1 d -allulose in 3 h. Key points • The present study explored an extreme temperature metagenome to identify a novel gene that encodes a thermostable l-rhamnose isomerase (L-RI M ) • L-RI M exhibits substantial (80% or more) activity in a broad spectrum of pH (6.0 to 9.0) and temperature (70 to 80 °C) ranges • L-RI M is excessively heat stable, displaying a half-life of about 12 days and 5 days at 65 °C and 70 °C, respectively
Sharma et al. (Tue,) studied this question.