Unexpected mode of action of sweet potato β-amylase on maltooligomer substrates

Erika Fazekas, Katalin Szabó, L. Kandra, G. Gyémánt

Research output: Article

10 Citations (Scopus)

Abstract

β-Amylase (EC 3.2.1.2), one of the main protein of the sweet potato, is an exo-working enzyme catalyzing the hydrolysis of α(1,4) glycosidic linkages in polysaccharides and removes successively maltose units from the non-reducing ends. The enzyme belongs to glycoside hydrolase GH14 family and inverts the anomeric configuration of the hydrolysis product. Multiple attack or processivity is an important property of polymer active enzymes and there is still limited information about the processivity of carbohydrate active enzymes. Action pattern and kinetic measurements of sweet potato β-amylase were made on a series of aromatic chromophor group-containing substrates (degree of polymerization DP 3-13) using HPLC method. Measured catalytic efficiencies increased with increasing DP of the substrates. Processive cleavage was observed on all substrates except the shortest pentamer. The mean number of steps without dissociation of enzyme-product complex increases with DP of substrate and reached 3.3 in case of CNPG11 indicating that processivity on longer substrates was more significant. A unique transglycosylation was observed on those substrates, which suffer processive cleavage and the substrates were re-built by the enzyme. Our results are the first presentation of a transglycosylation during an inverting glycosidase catalyzed hydrolysis. The yield of transglycosylation was remarkable high as shown in the change of the CNPG11 quantity. The CNPG11 concentration was doubled (from 0.24 to 0.54 mM) in the early phase of the reaction.

Original languageEnglish
Pages (from-to)1976-1981
Number of pages6
JournalBiochimica et Biophysica Acta - Proteins and Proteomics
Volume1834
Issue number10
DOIs
Publication statusPublished - 2013

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Ipomoea batatas
Amylases
Substrates
Enzymes
Hydrolysis
Glycoside Hydrolases
Maltose
Polymerization
Polysaccharides
Polymers
High Pressure Liquid Chromatography
Carbohydrates
Kinetics
Proteins

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Analytical Chemistry
  • Molecular Biology

Cite this

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title = "Unexpected mode of action of sweet potato β-amylase on maltooligomer substrates",
abstract = "β-Amylase (EC 3.2.1.2), one of the main protein of the sweet potato, is an exo-working enzyme catalyzing the hydrolysis of α(1,4) glycosidic linkages in polysaccharides and removes successively maltose units from the non-reducing ends. The enzyme belongs to glycoside hydrolase GH14 family and inverts the anomeric configuration of the hydrolysis product. Multiple attack or processivity is an important property of polymer active enzymes and there is still limited information about the processivity of carbohydrate active enzymes. Action pattern and kinetic measurements of sweet potato β-amylase were made on a series of aromatic chromophor group-containing substrates (degree of polymerization DP 3-13) using HPLC method. Measured catalytic efficiencies increased with increasing DP of the substrates. Processive cleavage was observed on all substrates except the shortest pentamer. The mean number of steps without dissociation of enzyme-product complex increases with DP of substrate and reached 3.3 in case of CNPG11 indicating that processivity on longer substrates was more significant. A unique transglycosylation was observed on those substrates, which suffer processive cleavage and the substrates were re-built by the enzyme. Our results are the first presentation of a transglycosylation during an inverting glycosidase catalyzed hydrolysis. The yield of transglycosylation was remarkable high as shown in the change of the CNPG11 quantity. The CNPG11 concentration was doubled (from 0.24 to 0.54 mM) in the early phase of the reaction.",
keywords = "β-Amylase, HPLC action pattern, Maltooligomers, Processive action, Sweet potato, Transglycosylation",
author = "Erika Fazekas and Katalin Szab{\'o} and L. Kandra and G. Gy{\'e}m{\'a}nt",
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TY - JOUR

T1 - Unexpected mode of action of sweet potato β-amylase on maltooligomer substrates

AU - Fazekas, Erika

AU - Szabó, Katalin

AU - Kandra, L.

AU - Gyémánt, G.

PY - 2013

Y1 - 2013

N2 - β-Amylase (EC 3.2.1.2), one of the main protein of the sweet potato, is an exo-working enzyme catalyzing the hydrolysis of α(1,4) glycosidic linkages in polysaccharides and removes successively maltose units from the non-reducing ends. The enzyme belongs to glycoside hydrolase GH14 family and inverts the anomeric configuration of the hydrolysis product. Multiple attack or processivity is an important property of polymer active enzymes and there is still limited information about the processivity of carbohydrate active enzymes. Action pattern and kinetic measurements of sweet potato β-amylase were made on a series of aromatic chromophor group-containing substrates (degree of polymerization DP 3-13) using HPLC method. Measured catalytic efficiencies increased with increasing DP of the substrates. Processive cleavage was observed on all substrates except the shortest pentamer. The mean number of steps without dissociation of enzyme-product complex increases with DP of substrate and reached 3.3 in case of CNPG11 indicating that processivity on longer substrates was more significant. A unique transglycosylation was observed on those substrates, which suffer processive cleavage and the substrates were re-built by the enzyme. Our results are the first presentation of a transglycosylation during an inverting glycosidase catalyzed hydrolysis. The yield of transglycosylation was remarkable high as shown in the change of the CNPG11 quantity. The CNPG11 concentration was doubled (from 0.24 to 0.54 mM) in the early phase of the reaction.

AB - β-Amylase (EC 3.2.1.2), one of the main protein of the sweet potato, is an exo-working enzyme catalyzing the hydrolysis of α(1,4) glycosidic linkages in polysaccharides and removes successively maltose units from the non-reducing ends. The enzyme belongs to glycoside hydrolase GH14 family and inverts the anomeric configuration of the hydrolysis product. Multiple attack or processivity is an important property of polymer active enzymes and there is still limited information about the processivity of carbohydrate active enzymes. Action pattern and kinetic measurements of sweet potato β-amylase were made on a series of aromatic chromophor group-containing substrates (degree of polymerization DP 3-13) using HPLC method. Measured catalytic efficiencies increased with increasing DP of the substrates. Processive cleavage was observed on all substrates except the shortest pentamer. The mean number of steps without dissociation of enzyme-product complex increases with DP of substrate and reached 3.3 in case of CNPG11 indicating that processivity on longer substrates was more significant. A unique transglycosylation was observed on those substrates, which suffer processive cleavage and the substrates were re-built by the enzyme. Our results are the first presentation of a transglycosylation during an inverting glycosidase catalyzed hydrolysis. The yield of transglycosylation was remarkable high as shown in the change of the CNPG11 quantity. The CNPG11 concentration was doubled (from 0.24 to 0.54 mM) in the early phase of the reaction.

KW - β-Amylase

KW - HPLC action pattern

KW - Maltooligomers

KW - Processive action

KW - Sweet potato

KW - Transglycosylation

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VL - 1834

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JO - Biochimica et Biophysica Acta - Proteins and Proteomics

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