Characterization of thermostable α-glucosidases from newly isolated Geobacillus sp. A333 and thermophilic bacterium A343

Abstract

We have partially purified and characterized two new thermostable exo-a-1,4-glucosidases (E.C.3.2.1.20) isolated from Geobacillus sp. A333 and thermophilic bacterium A343 strains. A333 a-glucosidase showed optimum activity at 60°C, pH 6.8 and had a value of 1.38 K m for the pNPG substrate, whereas these results were found to be 65°C, 7.0 and 0.85, respectively for A343 enzyme. Specificity for 20 different substrates and thin layer chromatography studies demonstrated that the A333 enzyme had high transglycosylation activity, and A343 had wide substrate specificity. The substrate specificity of A333 a-glucosidase was determined as maltose, dextrin, turanose, maltotriose, maltopentaose, meltotetraose, maltohexaose and phenyla-D-glycopyranoside. On the other hand, the A343 a-glucosidase mostly hydrolyzed dextrin, turanose, maltose, phenyl-a-D-glucopyranoside, maltotriose, maltotetraose, maltopentaose, isomaltose, saccharose and kojibiose by acting a-1,2, a-1,3, a-1,4 and a-1,6 bonds of these substrates. The relative activites of A333 and A343 enzymes were determined to be 83 and 92% when incubated at 60°C for 5 h whereas, the pH of 50% inactivation at 60°C for 15 h were determined to be pH 4.5/10.0 and pH 5.0/10.0, respectively. In addition, the results not only showed that both of the a-glucosidases were stable in a wide range of pH and temperatures, but were also found to be resistant to most of the denaturing agents, inhibitors and metal ions tested. With this study, thermostable exo-a-1,4-glucosidases produced by two new thermophilic strains were characterized as having biotechnological potential in transglycosylation reactions and starch hydrolysis processes.

Anoxybacillus salavatliensis sp. nov., an α‐glucosidase producing, thermophilic bacterium isolated from Salavatli, Turkey

“…T differs from the mentioned type strain by its ability of producing an intracellular, thermostable exo-α-1,4-glucosidase, having a specific activity of 0.67 U/mg with an optimum activity at 65 °C, pH 7.0 during 4 h of cultivation [16]. In addition, this strain is unique in that its α-glucosidase had a broad substrate specificity by acting α-1,2, α-1,3, α-1,4 and α-1,6 bonds of the substrates [16].…”

“…The thermostable exo-α-1,4-glucosidase of this strain, having biotechnological potential in starch hydrolysis processes, was also purified and characterized [16]. The current paper describes the taxonomy of the α-glucosidase producing strain A343…”

Geobacillus thermodenitrificans subsp. calidus, subsp. nov., a thermophilic and α-glucosidase producing bacterium isolated from Kizilcahamam, Turkey

“…The centrifuged supernatant was fractionated with 40 65% ammonium sulfate and dialyzed overnight and the 310 ml dialyzate was concentrated 20-fold by centrifugation in a speed vacuum concentrator. Then it was applied to a cation-exchange chromatography column as described previously (Cihan et al, 2009). Protein concentration was determined by using Bradford reagent, and as the protein standard 0.1 to 10 mg/ml concentrations of BSA were used (Cihan et al, 2009).…”

“…Then it was applied to a cation-exchange chromatography column as described previously (Cihan et al, 2009). Protein concentration was determined by using Bradford reagent, and as the protein standard 0.1 to 10 mg/ml concentrations of BSA were used (Cihan et al, 2009). When determining the molecular weight of the enzyme, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was carried out according to the procedure of Laemmli (1970).…”

“…Strain F84b T , F84a and A333 could produce starchdegrading enzymes with strong amylolytic and glycosidic activity, and α-glucosidase of strain A333 was characterized previously (Cihan et al, 2009). The α-glucosidase of strain A333 was found to be 45 kDa, having optimum activity at 60 C, and pH 6.8 in 16 h of cultivation.…”