Abstract:An alpha-glucosidase with the ability to attack polymeric substrates was purified to homogeneity from culture supernatants of Thermoanaerobacter thermohydrosulfuricus DSM 567. The enzyme is apparently a glycoprotein with a molecular mass of 160 kDa. Maximal activity is observed between pH5 and 7 at 75 degrees C. The alpha-glucosidase is active towards p-nitrophenyl-alpha-D-glucoside, maltose, malto-oligosaccharides, starch and pullulan. Highest activity is displayed towards the disaccharide maltose. In additio… Show more
“…The data obtained showed that we could assay the activity using maltose concentrations ranging from 5 to 8 mM. Various maltose concentrations including 3.3 mM (Wang and Hartman 1976 ), 5 mM (Bailey and Howard 1963 ), and 0.5% w/v (Wimmer et al 1997 ) have been used to assay maltase activity. A linear relationship between the amount of soil and the amount of glucose released (Fig.…”
Section: Resultsmentioning
confidence: 99%
“…6 ) suggesting the enzyme in soil is fairly stable. The optimum temperature for maltase extracted from soil microbes has been reported at 45 °C (Wang and Hartman 1976 ), 65 °C (Kobayashi et al 2003 ), 70 °C (Gomes et al 2005 ), and 75 °C (Wimmer et al 1997 ). Fig.…”
The enzyme maltase (glucoinvertase; glucosidosucrase; maltase-glucoamylase; α-glucopyranosidase; glucosidoinvertase; α-
d
-glucosidase; α-glucoside hydrolase; α-1,4-glucosidase EC 3.2.1.20), is involved in the exo-hydrolysis of 1,4-α-glucosidic linkages and certain oligosaccharides into glucose which is an important energy source for soil microbes. This enzyme originates from different sources, which include plants, seaweeds, protozoa, fungi, bacteria, vertebrates, and invertebrates. The assay of soil maltase using maltose as substrate and the released glucose determined using a glucose oxidase–peroxidase system has not been explored or investigated to the best of our knowledge. A simple assay protocol using this system is proposed to evaluate and characterize maltase activity in soils. The protocol involves the release of glucose (determined using a glucose oxidase–peroxidase colorimetric approach) when 1 g soil is treated with toluene and incubated with 5 mM maltose in 67 mM sodium acetate buffer (pH 5.0) at 37 °C for 1 h. The optimal activity using this procedure was at pH 5.0 and decreased at temperatures above 70 °C. The calculated
K
m
values ranged from 0.8 to 6.5 mM, and are comparable to those of enzymes purified from microorganisms. The Arrhenius equation plots for the activity in the four soils were linear between 20 and 70 °C. The activation energy values ranged from 34.1 to 57.2 kJ mol
−1
, the temperature coefficients (
Q
10
) ranged from 1.5 to 1.9 (avg. = 1.7), and the coefficients of variation (CV) of the proposed assay protocol for the soils used was <6%. While we recognize the availability of established assay protocols to determine soil α-glucosidase (referred in other literature as maltase) activity based on the
p
-nitrophenol (artificial product) released from
p
-nitrophenyl-α-
d
-glucopyranoside (artificial substrate), our interest was to assay its activity by determining the glucose (natural product) released from maltose (natural substrate).
“…The data obtained showed that we could assay the activity using maltose concentrations ranging from 5 to 8 mM. Various maltose concentrations including 3.3 mM (Wang and Hartman 1976 ), 5 mM (Bailey and Howard 1963 ), and 0.5% w/v (Wimmer et al 1997 ) have been used to assay maltase activity. A linear relationship between the amount of soil and the amount of glucose released (Fig.…”
Section: Resultsmentioning
confidence: 99%
“…6 ) suggesting the enzyme in soil is fairly stable. The optimum temperature for maltase extracted from soil microbes has been reported at 45 °C (Wang and Hartman 1976 ), 65 °C (Kobayashi et al 2003 ), 70 °C (Gomes et al 2005 ), and 75 °C (Wimmer et al 1997 ). Fig.…”
The enzyme maltase (glucoinvertase; glucosidosucrase; maltase-glucoamylase; α-glucopyranosidase; glucosidoinvertase; α-
d
-glucosidase; α-glucoside hydrolase; α-1,4-glucosidase EC 3.2.1.20), is involved in the exo-hydrolysis of 1,4-α-glucosidic linkages and certain oligosaccharides into glucose which is an important energy source for soil microbes. This enzyme originates from different sources, which include plants, seaweeds, protozoa, fungi, bacteria, vertebrates, and invertebrates. The assay of soil maltase using maltose as substrate and the released glucose determined using a glucose oxidase–peroxidase system has not been explored or investigated to the best of our knowledge. A simple assay protocol using this system is proposed to evaluate and characterize maltase activity in soils. The protocol involves the release of glucose (determined using a glucose oxidase–peroxidase colorimetric approach) when 1 g soil is treated with toluene and incubated with 5 mM maltose in 67 mM sodium acetate buffer (pH 5.0) at 37 °C for 1 h. The optimal activity using this procedure was at pH 5.0 and decreased at temperatures above 70 °C. The calculated
K
m
values ranged from 0.8 to 6.5 mM, and are comparable to those of enzymes purified from microorganisms. The Arrhenius equation plots for the activity in the four soils were linear between 20 and 70 °C. The activation energy values ranged from 34.1 to 57.2 kJ mol
−1
, the temperature coefficients (
Q
10
) ranged from 1.5 to 1.9 (avg. = 1.7), and the coefficients of variation (CV) of the proposed assay protocol for the soils used was <6%. While we recognize the availability of established assay protocols to determine soil α-glucosidase (referred in other literature as maltase) activity based on the
p
-nitrophenol (artificial product) released from
p
-nitrophenyl-α-
d
-glucopyranoside (artificial substrate), our interest was to assay its activity by determining the glucose (natural product) released from maltose (natural substrate).
“…The fact that T. vaginalis can use either of these polymers as carbon and energy sources suggests that additional α‐glucosidases must be responsible for their hydrolysis. A model for hydrolysis of starch proposed by Wimmer and coworkers predicts that the primary product will be maltohexaose or maltoheptaose (Wimmer et al 1997). If such an enzyme exists in T. vaginalis , it must have very low activity on pNGP, as it was not detected with this substrate.…”
Two isoforms of alpha-glucosidase were purified from the parasitic protist Trichomonas vaginalis. Both consisted of 103 kDa subunits, but differed in pH optimum and substrate specificity. Isoform 1 had a pH optimum around 4.5 and negligible activity on glucose oligomers other than maltose, while isoform 2 with a pH optimum of 5.5 hydrolyzed also such substrates at considerable rates. Neither had activity on glycogen or starch. Isoform 1 had a specific activity for hydrolysis of maltose of 30 U/mg protein and isoform 2 101 U/mg protein. The Km values were 0.4 mM and 2.0 mM, respectively. Isoform 2 probably corresponds to the activity detected on the cell surface.
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