ABSTRACT
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.
Subject(s)
Archaea/enzymology , Archaeal Proteins/chemistry , beta-Galactosidase/chemistry , Archaea/chemistry , Archaea/genetics , Archaeal Proteins/genetics , Archaeal Proteins/metabolism , Enzyme Stability , Galactose/metabolism , Glucosides/metabolism , Hot Temperature , Hydrogen-Ion Concentration , Kinetics , Lactose/metabolism , Substrate Specificity , beta-Galactosidase/genetics , beta-Galactosidase/metabolismABSTRACT
The aims of the current study were to evaluate the best technique for total phenolic extraction from Lavandula pubescens (Lp) and its application in vegetable oil industries as alternatives of synthetic food additives (TBHQ and BHT). To achieve these aims, three techniques of extraction were used: ultrasonic-microwave (40 kHz, 50 W, microwave power 480 W, 5 min), ultrasonic-homogenizer (20 kHz, 150 W, 5 min) and conventional maceration as a control. By using the Folin-Ciocalteu method, the total phenolic contents (TPC) (mg gallic acid equivalent/g dry matter) were found to be 253.87, 216.96 and 203.41 for ultrasonic-microwave extract, ultrasonic-homogenizer extract and maceration extract, respectively. The ultrasonic-microwave extract achieved the higher scavenger effect of DPPH (90.53%) with EC50 (19.54 µg/mL), and higher inhibition of ß-carotene/linoleate emulsion deterioration (94.44%) with IC50 (30.62 µg/mL). The activity of the ultrasonic-microwave treatment could prolong the induction period (18.82 h) and oxidative stability index (1.67) of fresh refined, bleached and deodorized palm olein oil (RBDPOo) according to Rancimat assay. There was an important synergist effect between citric acid and Lp extracts in improving the oxidative stability of fresh RBDPOo. The results of this work also showed that the ultrasonic-microwave assisted extract was the most effective against Gram-positive and Gram-negative strains that were assessed in this study. The uses of ultrasonic-microwave could induce the acoustic cavitation and rupture of plant cells, and this facilitates the flow of solvent into the plant cells and enhances the desorption from the matrix of solid samples, and thus would enhance the efficiency of extraction based on cavitation phenomenon.
