Studies on carbohydrate-metabolizing enzymes. 12. A survey of the carbohydrases of alfalfa

Manners, D. J.

doi:10.1042/bj0930545

Cited by 11 publications

(3 citation statements)

References 18 publications

(17 reference statements)

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“…Hydrolytic activity on raffinose and maltose but not on sucrose at pH 4.5 indicates the absence of an acid invertase but the presence of an ac-galactosidase and an a-glucosidase (Table I) but activities of this enzyme in other plants show pH optima of 3.5 to 5.5 (5,11). Acid a-glucosidase activity has been reported previously in sweet corn (12) and alfalfa (8). The occurrence of an acid a-glucosidase in sugar beet root is surprising but may indicate a genetic carryover from the starch-storing ancestors of the sugar beet.…”

Section: Resultsmentioning

confidence: 87%

Enzymes Involved in the Postharvest Degradation of Sucrose in Beta vulgaris L. Root Tissue

Wyse¹

1974

Plant Physiol.

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The reducing sugar content of sugar beet (Beta vulgaris L.) roots increased during 30 days of storage at 21 C and 160 days at 5 C as a result of an increase in acid invertase activity. Sucrose synthetase and neutral invertase activities were high at harvest but declined during storage, thus showing no relationship with postharvest reducing sugar accumulation in sugar beet roots. Acid a-glucosidase activity was detected in fresh roots but showed no activity with sucrose as a substrate.The sucrose content of sugar beet roots declines after harvest as a result of sucrose conversion to reducing sugars and oligosaccharides (20). At storage temperatures above 5 C reducing sugars (glucose and fructose) accumulate, while at temperatures below 5 C both raffinose and reducing sugars accumulate (20). The objective of this study was to determine which enzymes are involved in sucrose degradation in sugar beet roots during storage.Acid invertase activity is normally very low in fresh sugar beet roots, but induction of invertase activity by aging beet disks in water has been well documented (1,15,19 operations were carried out at 0 to 5 C. Roots were peeled, diced, and homogenized in a VirTis homogenizer' with an equal w/v ratio of grinding medium (0.05 M potassium phosphate, pH 7.2; 0.1 mm EDTA; 1 mM 2-mercaptoethanol; 10 mM Na2SO.). The homogenate was strained through two layers of dacron ninon and centrifuged at 16,000g for 20 min. The supernatant liquid was dialyzed overnight against 10 volumes of either grinding medium for neutral invertase and sucrose synthetase assays, or acetate buffer (0.05 M acetate, pH 5.0; 0.1 mM EDTA; 1 mm 2-mercaptoethanol; 10 mm Na,SO8) for acid invertase assay. The extract dialyzed at pH 5 was centrifuged at 1 6,000g for 10 min before assay and the pellet discarded.Assay. The standard assay mixture for neutral invertase contained 0.16 ml of buffer (8 ltmoles of potassium phosphate, pH 7.2; 0.016 lumole EDTA; 80 jumoles of sucrose) and 0.04 ml of the enzyme preparation. Normal assay time was 30 min at 35 C. Assay mixtures for sucrose synthetase were the same as for neutral invertase except that 10 ,u1 of 0.05 M UDP were added and only 0.03 ml of the enzyme preparation was used. Assay mixtures for acid invertase were the same as for neutral invertase except that the phosphate buffer was replaced by acetate at pH 5.0. The reactions were stopped by adding 1 ml of Somogyi's (18) copper reagent and reducing sugars determined by the method of Nelson (14). Glucose liberation, in assays in which maltose was the substrate, was determined by the glucose oxidase/peroxidase method with a commercial reagent (Glucostat, Worthington2).Protein was determined by the method of Lowry (9). The reducing sugar content of the stored roots was determined on a clarified extract prepared by the method of Dexter et al. (4). RESULTS AND DISCUSSIONIn a preliminary experiment a survey of enzymes which may degrade sucrose in fresh sugar beet roots was made. Sucrose, raffinose, and maltose were used as substrates and a...

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Section: Resultsmentioning

confidence: 87%

Enzymes Involved in the Postharvest Degradation of Sucrose in Beta vulgaris L. Root Tissue

Wyse¹

1974

Plant Physiol.

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“…The hydrolytic activity of the seedling extracts is due to plant enzymes, and not to contaminating bacterial enzymes, since control experiments have shown that the activity of extracts of alfalfa seeds germinated under sterile conditions was identical with that of seeds germinated under the usual conditions (Hutson & Manners, 1964).…”

Section: Discussionmentioning

confidence: 99%

“…Enzymes which hydrolyse oa-glucosidic linkages are widely distributed in Nature; for example, extracts of various land plants, seaweeds, protozoa, fungi, bacteria, vertebrates and invertebrates contain maltase (for a review see Larner, 1960). During a survey of the carbohydrase activity of alfalfa (lucerne) extracts (Hutson & Manners, 1964), strong maltase activity was observed; however, the degradation of soluble nigeran [a linear polymer of D-glucopyranose containing alternate a-( 1-4)and oc-(1-3)-glucosidic linkages which is resistant to a-and fi-amylolysis] to glucose by the extracts was an unexpected feature. This breakdown indicated that one or more oa-(1-*3)-glucosidases were present in the extracts.…”

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confidence: 99%

STUDIES ON CARBOHYDRATE-METABOLIZING ENZYMES. THE HYDROLYSIS OF α-GLUCOSIDES, INCLUDING NIGEROSE, BY EXTRACTS OF ALFALFA AND OTHER HIGHER PLANTS

Manners¹

1965

Biochemical Journal

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1. Enzyme preparations from 11 plant sources, from yeast and from the protozoan Tetrahymena pyriformis show nigerase activity, which, in most preparations, was 70-90% of that towards maltose. 2. These enzyme preparations also hydrolysed isomaltose, but there was a wide variation in relative maltase to isomaltase activity. 3. The maltase and nigerase activities of alfalfa and tomato preparations could not be differentiated by heat inactivation or inhibitor methods. However, with turanose used as a competitive inhibitor, evidence suggesting that maltose and nigerose are hydrolysed at different catalytically active sites in the alfalfa preparation was obtained. 4. It is probable that the alfalfa alpha-glucosidase exists as a mixture of isoenzymes.

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The Biochemistry of Laminarin and the Nature of Laminarinase

Bull¹,

Chesters²

1966

Advances in Enzymology - And Related Areas of Molecular Biology

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Studies on carbohydrate-metabolizing enzymes. 12. A survey of the carbohydrases of alfalfa

Cited by 11 publications

References 18 publications

Enzymes Involved in the Postharvest Degradation of Sucrose in Beta vulgaris L. Root Tissue

Enzymes Involved in the Postharvest Degradation of Sucrose in Beta vulgaris L. Root Tissue

STUDIES ON CARBOHYDRATE-METABOLIZING ENZYMES. THE HYDROLYSIS OF α-GLUCOSIDES, INCLUDING NIGEROSE, BY EXTRACTS OF ALFALFA AND OTHER HIGHER PLANTS

The Biochemistry of Laminarin and the Nature of Laminarinase

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