β-Galactosidase from Exiguobacterium acetylicum: Cloning, expression, purification and characterization

Aburto, Carla; Castillo, Carlos; Cornejo, Fabián A.; Arenas-Salinas, Mauricio; Vásquez, Claudio C.; Guerrero, Cecilia; Arenas, Felipe; Illanes, Andrés; Vera, Carlos

doi:10.1016/j.biortech.2019.01.005

Cited by 19 publications

(6 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The activity of β-galactosidases (BLGLB1 and BPGLB1) was measured by using oNPG (2 mM) dissolved in a phosphate buffer solution (pH 7.0, 10 mM) as the substrate [ 6 ]. One international activity unit was defined as the amount of β-galactosidase that hydrolyzed 1 μmoL oNPG per minute.…”

Section: Methodsmentioning

confidence: 99%

“…β-galactosidase with trans glycosylation activity participates in the transglycosylation reaction to produce prebiotic galactooligosaccharides [ 1 , 2 ]. In addition, β-galactosidase can also be used as a biocatalyst to hydrolyze lactose, which is widely used in dairy processing, food processing, and pharmaceutical fields products [ 3 , 4 , 5 , 6 ]. For example, whey is a byproduct of cheese production and contains 4–4.5% lactose.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Cloning, Expression, Purification, and Characterization of β-Galactosidase from Bifidobacterium longum and Bifidobacterium pseudocatenulatum

Yang

Jiang

et al. 2022

Molecules

View full text Add to dashboard Cite

Expression and purification of β-galactosidases derived from Bifidobacterium provide a new resource for efficient lactose hydrolysis and lactose intolerance alleviation. Here, we cloned and expressed two β-galactosidases derived from Bifidobacterium. The optimal pH for BLGLB1 was 5.5, and the optimal temperature was 45 °C, at which the enzyme activity of BLGLB1 was higher than that of commercial enzyme E (300 ± 3.6 U/mg) under its optimal conditions, reaching 2200 ± 15 U/mg. The optimal pH and temperature for BPGLB1 were 6.0 and 45 °C, respectively, and the enzyme activity (0.58 ± 0.03 U/mg) under optimum conditions was significantly lower than that of BLGLB1. The structures of the two β-galactosidase were similar, with all known key sites conserved. When o-nitrophenyl-β-D-galactoside (oNPG) was used as an enzyme reaction substrate, the maximum reaction velocity (Vmax) for BLGLB1 and BPGLB1 was 3700 ± 100 U/mg and 1.1 ± 0.1 U/mg, respectively. The kinetic constant (Km) of BLGLB1 and BPGLB1 was 1.9 ± 0.1 and 1.3 ± 0.3 mmol/L, respectively. The respective catalytic constant (kcat) of BLGLB1 and BPGLB1 was 1700 ± 40 s−1 and 0.5 ± 0.02 s−1, respectively; the respective kcat/Km value of BLGLB1 and BPGLB1 was 870 L/(mmol∙s) and 0.36 L/(mmol∙s), respectively. The Km, kcat and Vmax values of BLGLB1 were superior to those of earlier reported β-galactosidase derived from Bifidobacterium. Overall, BLGLB1 has potential application in the food industry.

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Cloning, Expression, Purification, and Characterization of β-Galactosidase from Bifidobacterium longum and Bifidobacterium pseudocatenulatum

Yang

Jiang

et al. 2022

Molecules

View full text Add to dashboard Cite

show abstract

“…Furthermore, this enzyme was stable at pH ranges of 5 to 8, and retained more than 75% of its original activity after 2 h of incubation (Figure 3C). Most reported optimal pH for the enzymatic activity of GH42 β-galactosidase was in the pH range of 6 to 7, 10,17,37,38 except for B. coagulans β-galactosidase BcGalB and P. torridus β-galactosidase PTO1453, with the optimal pH values of 5.0 and 5.5, 28,39 respectively. BtGal42 displayed the maximum activity at 50 • C, which was quite similar to that of β-galactosidases from P. marinomonas ef1, 40 B. velezensis SW5, 41 L. culinaris, 42 and B. adolescentis DSM20083, 43 while was higher than that of β-galactosidases from B. adolescentis BgaC, 29 B. bifidum (Saphera), 32 K. oxytoca ZJUH1705, 17 and B. bifidum S17.…”

Section: Biochemical Properties Of Btgal42mentioning

confidence: 99%

A GH42 β‐galactosidase from Bifidobacterium thermophilum: Biochemical characterization and synthesis of galactosyl glycerol by reverse hydrolysis

Duan

Jia

Cao

et al. 2022

Biotech and App Biochem

View full text Add to dashboard Cite

In this study, a β‐galactosidase gene (btgal42) was first cloned from Bifidobacterium thermophilum and successfully expressed in Escherichia coli. The molecular weight of the purified BtGal42 was estimated to be 78 kDa by the sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and 225 kDa by the size‐exclusion chromatography, indicating that the native BtGal42 was a homotrimer. BtGal42 belonged to the glycosidase hydrolase family 42, exhibiting the maximum activity at pH of 7 and at temperature of 50°C. The enzyme displayed a strictly specific activity toward substrates with β‐galactosyl linkages at the nonreducing ends, of which the activity on 4‐nitrophenyl‐β‐d‐galactopyranoside was the highest, followed by 2‐nitrophenyl‐β‐d‐galactopyranoside and lactose. Among the tested metals and reagents, BtGal42 showed tolerance in the presence of various organic solvents. Importantly, BtGal42 exhibited a high reverse hydrolysis activity when using galactose as the donor and the di‐alcohol ethylene glycol and the trialcohol glycerol as the acceptors. Under unoptimized reaction conditions, the galactosyl glycerol yield reached 62.2 g/L (galactose conversion rate, 41.2%). This study might provide a feasible method for the biosynthesis of galactosyl glycerol from low‐cost glycerol and galactose, which was associated with high conversion efficiency and few byproducts.

show abstract

“…ON14, isolated from Antarctica, exhibits high catalytic activity at 15 • C [18]. β-Galactosidase from Exiguobacterium acetylicum can maintain 80% residual enzyme activity at 40 • C for 200 h and has good thermostability [19]. Furthermore, the structures of GH-42 β-galactosidases have also been described in theoretical studies.…”

Section: Introductionmentioning

confidence: 99%

A Novel Thermal-Activated β-Galactosidase from Bacillus aryabhattai GEL-09 for Lactose Hydrolysis in Milk

Luan

Duan

2022

Foods

View full text Add to dashboard Cite

β-Galactosidase has been greatly used in the dairy industry. This study investigated a novel thermostable β-galactosidase (lacZBa) from Bacillus aryabhattai GEL-09 and evaluated the hydrolytic performance of this enzyme. Firstly, the lacZBa-encoding gene was cloned and overexpressed in Escherichia coli BL21(DE3). Phylogenetic analyses revealed that lacZBa belonged to the glycoside hydrolase family 42. Using SDS-PAGE, we determined that the molecular weight of lacZBa was ~75 kDa. Purified lacZBa exhibited a maximum activity at 45 °C, pH 6.0, and could be activated following incubation at 45 °C for several minutes. The half-life of lacZBa at 45 °C and 50 °C was 264 h and 36 h, respectively. While Co2+, Mn2+, Zn2+, Fe2+, Mg2+, and Ca2+ enhanced enzymatic activity, Cu2+ and ethylenediaminetetraacetic acid inhibited enzymatic activity. Moreover, lacZBa could hydrolyze lactose and oNPG with Km values of 85.09 and 14.38 mM. Molecular docking results revealed that lacZBa efficiently recognized and catalyzed lactose. Additionally, the hydrolysis of lactose by lacZBa was studied in lactose solution and commercial milk. Lactose was completely hydrolyzed within 4 h with 8 U/mL of lacZBa at 45 °C. These results suggested that lacZBa identified in this study has potential applications in the dairy industry.

show abstract

β-Galactosidase from Exiguobacterium acetylicum: Cloning, expression, purification and characterization

Cited by 19 publications

References 22 publications

Cloning, Expression, Purification, and Characterization of β-Galactosidase from Bifidobacterium longum and Bifidobacterium pseudocatenulatum

Cloning, Expression, Purification, and Characterization of β-Galactosidase from Bifidobacterium longum and Bifidobacterium pseudocatenulatum

A GH42 β‐galactosidase from Bifidobacterium thermophilum: Biochemical characterization and synthesis of galactosyl glycerol by reverse hydrolysis

A Novel Thermal-Activated β-Galactosidase from Bacillus aryabhattai GEL-09 for Lactose Hydrolysis in Milk

Contact Info

Product

Resources

About