Modulation of the interaction between aldolase and glycerol-phosphate dehydrogenase by fructose phosphates

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Abstract

Kinetics of fructose-1,6-disphosphate aldolase (EC 4.1.2.13) catalyzed conversion of fructose phosphates was analyzed by coupling the aldolase reactions to the metabolically sequential enzyme, glycerol-3-phosphate dehydrogenase (EC 1.1.1.8), which interacts with aldolase. At low enzyme concentration poly(ethylene glycol) was added to promote complex formation of aldolase and glycerol-phosphate dehydrogenase resulting in a 3-fold increase in KM of fructose-1,6-bisphosphate and no change in Vmax. Kinetic parameters for fructose-1-phosphate conversion changed inversely upon complex formation: Vmax increased while KM remained unchanged. Gel penetration and ion-exchange chromatographic experiments showed positive modulation of the interaction of aldolase and dehydrogenase by fructose-1,6-bisphosphate. The dissociation constant of the heterologous enzyme complex decreased 10-fold in the presence of this substrate. Fructose-1-phosphate or dihydroxyacetone phosphate had no effect on the dissociation constant of the aldolase-dehydrogenase complex. In addition, titration of fluorescein-labelled glycerolphosphate dehydrogenase with aldolase indicated that both fructose-1,6-bisphosphate and fructose-2,6-bisphosphate enhanced the affinity of aldolase to glycerol-phosphate dehydrogenase. The results of the kinetic and binding experiments suggest that binding of the C-6 phosphate group of fructose-1,6-bisphosphate to aldolase complexed with dehydrogenase is sterically impeded while saturation of the C-6 phosphate group site increases the affinity of aldolase for dehydrogenase. The possible molecular mechanism of the fructose-1,6-bisphosphate modulated interaction is discussed.

Original languageEnglish
Pages (from-to)236-242
Number of pages7
JournalBiochimica et Biophysica Acta (BBA)/Protein Structure and Molecular
Volume1078
Issue number2
DOIs
Publication statusPublished - Jun 24 1991

Fingerprint

glycerol dehydrogenase
Fructose-Bisphosphate Aldolase
Fructose
Glycerol
Oxidoreductases
Phosphates
Modulation
Glycerolphosphate Dehydrogenase
Enzymes
Dihydroxyacetone Phosphate
Kinetics
Ethylene Glycol
Ion Exchange

Keywords

  • Bienzyme complex
  • Dynamism
  • Fructose bisphosphate
  • Specificity
  • Substrate modulated interaction

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology
  • Structural Biology
  • Medicine(all)

Cite this

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title = "Modulation of the interaction between aldolase and glycerol-phosphate dehydrogenase by fructose phosphates",
abstract = "Kinetics of fructose-1,6-disphosphate aldolase (EC 4.1.2.13) catalyzed conversion of fructose phosphates was analyzed by coupling the aldolase reactions to the metabolically sequential enzyme, glycerol-3-phosphate dehydrogenase (EC 1.1.1.8), which interacts with aldolase. At low enzyme concentration poly(ethylene glycol) was added to promote complex formation of aldolase and glycerol-phosphate dehydrogenase resulting in a 3-fold increase in KM of fructose-1,6-bisphosphate and no change in Vmax. Kinetic parameters for fructose-1-phosphate conversion changed inversely upon complex formation: Vmax increased while KM remained unchanged. Gel penetration and ion-exchange chromatographic experiments showed positive modulation of the interaction of aldolase and dehydrogenase by fructose-1,6-bisphosphate. The dissociation constant of the heterologous enzyme complex decreased 10-fold in the presence of this substrate. Fructose-1-phosphate or dihydroxyacetone phosphate had no effect on the dissociation constant of the aldolase-dehydrogenase complex. In addition, titration of fluorescein-labelled glycerolphosphate dehydrogenase with aldolase indicated that both fructose-1,6-bisphosphate and fructose-2,6-bisphosphate enhanced the affinity of aldolase to glycerol-phosphate dehydrogenase. The results of the kinetic and binding experiments suggest that binding of the C-6 phosphate group of fructose-1,6-bisphosphate to aldolase complexed with dehydrogenase is sterically impeded while saturation of the C-6 phosphate group site increases the affinity of aldolase for dehydrogenase. The possible molecular mechanism of the fructose-1,6-bisphosphate modulated interaction is discussed.",
keywords = "Bienzyme complex, Dynamism, Fructose bisphosphate, Specificity, Substrate modulated interaction",
author = "B. V{\'e}rtessy and F. Orosz and J. Ov{\'a}di",
year = "1991",
month = "6",
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doi = "10.1016/0167-4838(91)90564-G",
language = "English",
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T1 - Modulation of the interaction between aldolase and glycerol-phosphate dehydrogenase by fructose phosphates

AU - Vértessy, B.

AU - Orosz, F.

AU - Ovádi, J.

PY - 1991/6/24

Y1 - 1991/6/24

N2 - Kinetics of fructose-1,6-disphosphate aldolase (EC 4.1.2.13) catalyzed conversion of fructose phosphates was analyzed by coupling the aldolase reactions to the metabolically sequential enzyme, glycerol-3-phosphate dehydrogenase (EC 1.1.1.8), which interacts with aldolase. At low enzyme concentration poly(ethylene glycol) was added to promote complex formation of aldolase and glycerol-phosphate dehydrogenase resulting in a 3-fold increase in KM of fructose-1,6-bisphosphate and no change in Vmax. Kinetic parameters for fructose-1-phosphate conversion changed inversely upon complex formation: Vmax increased while KM remained unchanged. Gel penetration and ion-exchange chromatographic experiments showed positive modulation of the interaction of aldolase and dehydrogenase by fructose-1,6-bisphosphate. The dissociation constant of the heterologous enzyme complex decreased 10-fold in the presence of this substrate. Fructose-1-phosphate or dihydroxyacetone phosphate had no effect on the dissociation constant of the aldolase-dehydrogenase complex. In addition, titration of fluorescein-labelled glycerolphosphate dehydrogenase with aldolase indicated that both fructose-1,6-bisphosphate and fructose-2,6-bisphosphate enhanced the affinity of aldolase to glycerol-phosphate dehydrogenase. The results of the kinetic and binding experiments suggest that binding of the C-6 phosphate group of fructose-1,6-bisphosphate to aldolase complexed with dehydrogenase is sterically impeded while saturation of the C-6 phosphate group site increases the affinity of aldolase for dehydrogenase. The possible molecular mechanism of the fructose-1,6-bisphosphate modulated interaction is discussed.

AB - Kinetics of fructose-1,6-disphosphate aldolase (EC 4.1.2.13) catalyzed conversion of fructose phosphates was analyzed by coupling the aldolase reactions to the metabolically sequential enzyme, glycerol-3-phosphate dehydrogenase (EC 1.1.1.8), which interacts with aldolase. At low enzyme concentration poly(ethylene glycol) was added to promote complex formation of aldolase and glycerol-phosphate dehydrogenase resulting in a 3-fold increase in KM of fructose-1,6-bisphosphate and no change in Vmax. Kinetic parameters for fructose-1-phosphate conversion changed inversely upon complex formation: Vmax increased while KM remained unchanged. Gel penetration and ion-exchange chromatographic experiments showed positive modulation of the interaction of aldolase and dehydrogenase by fructose-1,6-bisphosphate. The dissociation constant of the heterologous enzyme complex decreased 10-fold in the presence of this substrate. Fructose-1-phosphate or dihydroxyacetone phosphate had no effect on the dissociation constant of the aldolase-dehydrogenase complex. In addition, titration of fluorescein-labelled glycerolphosphate dehydrogenase with aldolase indicated that both fructose-1,6-bisphosphate and fructose-2,6-bisphosphate enhanced the affinity of aldolase to glycerol-phosphate dehydrogenase. The results of the kinetic and binding experiments suggest that binding of the C-6 phosphate group of fructose-1,6-bisphosphate to aldolase complexed with dehydrogenase is sterically impeded while saturation of the C-6 phosphate group site increases the affinity of aldolase for dehydrogenase. The possible molecular mechanism of the fructose-1,6-bisphosphate modulated interaction is discussed.

KW - Bienzyme complex

KW - Dynamism

KW - Fructose bisphosphate

KW - Specificity

KW - Substrate modulated interaction

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DO - 10.1016/0167-4838(91)90564-G

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AN - SCOPUS:0025735063

VL - 1078

SP - 236

EP - 242

JO - Biochimica et Biophysica Acta - Proteins and Proteomics

JF - Biochimica et Biophysica Acta - Proteins and Proteomics

SN - 1570-9639

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