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Characterization of L-rhamnose isomerase from Clostridium stercorarium and its application to the production of D-allose from D-allulose (D-psicose)

Seo, Min-Ju, Choi, Ji-Hyeon, Kang, Su-Hwan, Shin, Kyung-Chul, Oh, Deok-Kun
Biotechnology letters 2018 v.40 no.2 pp. 325-334
Clostridium stercorarium, allose, catalytic activity, half life, isomerases, isomerization, manganese, pH, psicose, rhamnose, temperature, thermophilic bacteria
OBJECTIVE: To characterize L-rhamnose isomerase (L-RI) from the thermophilic bacterium Clostridium stercorarium and apply it to produce D-allose from D-allulose. RESULTS: A recombinant L-RI from C. stercorarium exhibited the highest specific activity and catalytic efficiency (k cₐₜ/K ₘ) for L-rhamnose among the reported L-RIs. The L-RI was applied to the high-level production of D-allose from D-allulose. The isomerization activity for D-allulose was maximal at pH 7, 75 °C, and 1 mM Mn²⁺ over 10 min reaction time. The half-lives of the L-RI at 65, 70, 75, and 80 °C were 22.8, 9.5, 1.9, and 0.2 h, respectively. To ensure full stability during 2.5 h incubation, the optimal temperature was set at 70 °C. Under the optimized conditions of pH 7, 70 °C, 1 mM Mn²⁺, 27 U L-RI l⁻¹, and 600 g D-allulose l⁻¹, L-RI from C. stercorarium produced 199 g D-allose l⁻¹ without by-products over 2.5 h, with a conversion yield of 33% and a productivity of 79.6 g l⁻¹ h⁻¹. CONCLUSION: To the best of our knowledge, this is the highest concentration and productivity of D-allose reported thus far.