Lactulose production by a thermostable glycoside hydrolase from the hyperthermophilic archaeon Caldivirga maquilingensis IC-167.

BACKGROUND: Lactulose has various uses in the food and pharmaceutical fields. Thermostable enzymes have many advantages for industrial exploitation, including high substrate solubilities as well as reduced risk of process contamination. RESULTS: Enzymatic synthesis of lactulose employing a transgalactosylation reaction by a recombinant thermostable glycoside hydrolase (GH1) from the hyperthermophilic archaeon Caldivirga maquilingensis IC-167 was investigated. The optimal pH for lactulose production was found to be 4.5, while the optimal temperature was 85 °C, before it dropped moderately to 83% at 90 °C. However, the relative activity for lactulose synthesis dropped sharply to 35% at 95 °C. At optimal reaction conditions of 70% (w/w) initial sugar substrates with molar ratio of lactose to fructose of 1:4, 15 U mL-1 enzyme concentration and 85 °C, the time course reaction produced a maximum lactulose concentration of 108 g L-1 at 4 h, corresponding to a lactulose yield of 14% and 27 g L-1 h-1 productivity with 84% lactose conversion. The transgalactosylation reaction for lactulose synthesis was greatly influenced by the ratio of galactose donor to acceptor. CONCLUSION: This novel GH1 may be useful for process applications owing to its high activity in very concentrated substrate reaction media and promising thermostability. © 2017 Society of Chemical Industry.

Characterization of a thermostable glycoside hydrolase (CMbg0408) from the hyperthermophilic archaeon Caldivirga maquilingensis IC-167.

BACKGROUND: Hyperthermophilic archaea capable of functioning optimally at very high temperatures are a good source of unique and industrially important thermostable enzymes. RESULTS: A glycoside hydrolase family 1 ß-galactosidase gene (BglB) from a hyperthermophilic archaeon Caldivirga maquilingensis IC-167 was cloned and expressed in Escherichia coli. The recombinant enzyme (CMbg0408) displayed optimum activity at 110 °C and pH 5.0. It also retained 92% and 70% of its maximal activity at 115 and 120 °C, respectively. The enzyme was completely thermostable and active after 120 min of incubation at 80 and 90 °C. It also showed broad substrate specificity with activities of 8876 ± 185 U mg-1 for p-nitrophenyl-ß-d-galactopyranoside, 4464 ± 172 U mg-1 for p-nitrophenyl-ß-d-glucopyranoside, 1486 ± 68 U mg-1 for o-nitrophenyl-ß-d-galactopyranoside, 2250 ± 86 U mg-1 for o-nitrophenyl-ß-d-xylopyranoside and 175 ± 4 U mg-1 for lactose. A catalytic efficiency (kcat /Km ) of 3059 ± 122 mmol L-1 s-1 and Km value of 8.1 ± 0.08 mmol L-1 were displayed towards p-nitrophenyl-ß-d-galactopyranoside. CONCLUSION: As a result of its remarkable thermostability and high activity at high temperatures, this novel ß-galactosidase may be useful for food and pharmaceutical applications. © 2016 Society of Chemical Industry.