Gene clone and characterization of a novel thermostable β-galactosidase with transglycosylation activity from Thermotoga naphthophila RUK-10

Abstract

We cloned and expressed a new recombinant β-galactosidase(TN0949) from Thermotoga naphthophila RKU-10 with the pET28a(+) vector system in Escherichia coli BL21(DE3), and determined its catalytic capability to synthesize alkyl glucosides. The recombinant enzyme was purified to a single band via heat treatment and Ni 2+ -NTA affinity chromatography. The molecular mass of the recombinant enzyme was estimated to be 79 kDa with sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE). TN0949 can hydrolyze o-nitrophenyl β-Dgalactopyranoside at the optimum pH and temperature of 6.5 and 80 °C, respectively. TN0949 can also hydrolyze lactose at the optimum pH and temperature of 5.2 and 80 °C, respectively. The K m values for the hydrolyses of o-nitrophenyl β-D-galactopyranoside and lactose were 0.82 and 83.65 mmol/L, respectively. TN0949 was stable over a wide range of pH(3.0 to 7.0) after 24 h of incubation. The half-lives of TN0949 at 75, 80 and 85 °C were 22, 6 and 1.33 h, respectively. The enzyme displayed the capability to use lactose as the transglycosylation substrate to synthesize butyl galactopyranoside and hexyl galactopyranoside, indicating its suitability as a candidate industrial biocatalyst.

Synergistic effect of thermostable β-glucosidase TN0602 and cellulase on cellulose hydrolysis

“…This is also the rate-limiting step of the entire hydrolysis process. Therefore, b-glucosidases is a key factor that determines the efficiency of cellulose hydrolysis (Yang et al 2015a).…”

Purification and catalytic behavior optimization of lactose degrading β-galactosidase from Aspergillus nidulans

“…CQ31 showed 25.1 kDa molecular mass (Lee et al 2003;Jiang et al 2010). The molecular mass of the recombinant b-galactosidase from Thermotoga naphthophila was estimated to be 79.0 kDa (Yang et al 2015) however, the molecular mass of the bgalactosidase from Pediococcus acidilactici as measured by the SDS-PAGE and MALDI-TOF was 39.07 kDa (Chanalia et al 2018).…”