scholarly journals Comparison of the kinetic properties of the pyruvate dehydrogenase complex from pig kidney cortex and medulla.

1993 ◽  
Vol 40 (3) ◽  
pp. 411-419 ◽  
Author(s):  
T Pawełczyk ◽  
M S Olson
Keyword(s):  
Ionic Strength ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Complex ◽  
Product Inhibition ◽  
Hill Coefficient ◽  
Kidney Cortex ◽  
Kinetic Properties ◽  
Kidney Medulla ◽  
Pig Kidney ◽  
Dehydrogenase Complex

The activity of the pyruvate dehydrogenase complex (PDC) purified from pig kidney medulla was affected by K+, Na+, Cl-, HCO3-, HPO4(2-) and changes in ionic strength. Increased ionic strength influenced the activity of PDC from medulla by decreasing the Vmax and S0.5 for pyruvate and increasing the Hill coefficient. The magnitude of these changes was smaller than the corresponding changes for PDC purified from the cortex. In the presence of K+ (80 mM), Na+ (20 mM), Cl- (20 mM), HCO3- (20 mM), HPO4(2-) (10 mM) and at ionic strength of 0.15 M the S0.5 for pyruvate of PDC from medulla was 117 microM and the enzyme complex was saturated by 1.1 mM pyruvate. Under these conditions the S0.5 for pyruvate of PDC derived from cortex was 159 microM and the enzyme was saturated at 4.5 mM pyruvate. Based on the results presented in this report it is suggested that PDC in kidney medulla may be regulated not only by a phosphorylation/dephosphorylation system and end-product inhibition but also via changes in ionic strength.

scholarly journals The effects of various anions and cations on the regulation of pyruvate dehydrogenase complex activity from pig kidney cortex

1988 ◽  
Vol 253 (3) ◽  
pp. 819-825 ◽  
Author(s):  
T Pawelczyk ◽  
R A Easom ◽  
M S Olson
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Complex ◽  
Maximal Activity ◽  
Hill Coefficient ◽  
Kidney Cortex ◽  
Complex Activity ◽  
Pig Kidney ◽  
Constant Strength ◽  
Dehydrogenase Complex

The activity of pyruvate dehydrogenase complex (PDC) purified from pig kidney cortex was found to be affected by various uni- and bi-valent ions. At a constant strength of 0.13 M at pH 7.8, K+, Na+, Cl-, HCO3- and HPO4(2-) had significant effects on the activity of PDC: Na+, K+ and HPO4(2-) stimulated, but HCO3- and Cl- inhibited. The stimulatory effect of Na+ was mediated by a change in the Vmax. of PDC only, whereas K+ produced an increase in Vmax. and a change in the Hill coefficient (h). The extent of stimulation produced by HPO4(2-)4 on the activity of PDC was dependent on the concentrations of K+ and Na+. Both cations at concentrations higher than 40 mM partially prevented the effect of HPO4(2-)4. Cl- and HCO3- anions decreased the Vmax. of the enzyme and increased the S0.5 for pyruvate. The effects of Na+, K+, Cl-, HPO4(2-) and HCO3- on the activity of PDC were additive. In the presence of 80 mM-K+, 20 mM-Na+, 10 mM-HPO4(2-), 20 mM-Cl- and 20 mM-HCO3- the activity of PDC was increased by 30%, the S0.5 for pyruvate was increased from 75 to 158 microM and h was decreased from 1.3 to 1.1. Under these conditions and at 1.0 mM-pyruvate, the activity of PDC was 80% of the maximal activity achieved in the presence of these ions and 4.5 mM-pyruvate. The present study suggests that PDC may operate under non-saturating concentrations for substrate in vivo.

scholarly journals The dependence of electrophoretic and spectroscopic properties of the pyruvate dehydrogenase complex on mono- and divalent ions.

1994 ◽  
Vol 41 (1) ◽  
pp. 63-72
Author(s):  
T Pawełczyk ◽  
R A Easom ◽  
M S Olson
Keyword(s):  
Ionic Strength ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Complex ◽  
Fluorescence Emission ◽  
Emission Spectra ◽  
Spectroscopic Properties ◽  
Uv Absorption ◽  
Divalent Ions ◽  
Fluorescence Emission Spectra ◽  
Dehydrogenase Complex

The effects of various mono- and divalent ions on the pyruvate dehydrogenase complex (PDC) were investigated. To determine the radius of PDC under various conditions a two-dimensional agarose gel electrophoresis technique was used. The radius of PDC cross-linked with glutaraldehyde at ionic strength 0.04 M was calculated to be 22.0 +/- 0.1 nm. The presence of K+, Na+ or HPO4(2-) prevented changes in electromobility and of the calculated radius of PDC induced by alteration in ionic strength. The fluorescence emission spectra of PDC depended on the ionic strength and monovalent cations. The fluorescence intensity of PDC increased in the presence of 80 mM K+, and decreased in the presence of 80 mM Na+ with no shift in the emission maximum wavelength. Changes in the ionic strength to which PDC was exposed resulted in alteration of the UV absorption spectra in the 230 nm region. These alterations were prevented by HPO4(2-), whereas Na+ or K+ ions had no effect on the UV absorption spectrum of PDC.

scholarly journals The plasticity of the pyruvate dehydrogenase complex confers a labile structure that is associated with its catalytic activity

2020 ◽  
Author(s):  
Jaehyoun Lee ◽  
Seunghee Oh ◽  
Saikat Bhattacharya ◽  
Ying Zhang ◽  
Laurence Florens ◽  
...  
Keyword(s):  
Ionic Strength ◽  
Pyruvate Dehydrogenase ◽  
Biochemical Analysis ◽  
Pyruvate Dehydrogenase Complex ◽  
Size Exclusion ◽  
Dependent Manner ◽  
Acetyl Coa ◽  
Cell Extracts ◽  
Exclusion Chromatography ◽  
Dehydrogenase Complex

ABSTRACTThe pyruvate dehydrogenase complex (PDC) is a multienzyme complex that plays a key role in energy metabolism by converting pyruvate to acetyl-CoA. An increase of nuclear PDC has been shown to be correlated with an increase of histone acetylation that requires acetyl-CoA. PDC has been reported to form a ~ 10 MDa macromolecular machine that is proficient in performing sequential catalytic reactions via its three components. In this study, we show that the PDC displays size versatility in an ionic strength-dependent manner using size exclusion chromatography of yeast cell extracts. Biochemical analysis in combination with mass spectrometry indicates that yeast PDC (yPDC) is a salt-labile complex that dissociates into sub-megadalton individual components even under physiological ionic strength. Interestingly, we find that each oligomeric component of yPDC displays a larger size than previously believed. In addition, we show that the mammalian PDC also displays this uncommon characteristic of salt-lability, although it has a somewhat different profile compared to yeast. We show that the activity of yPDC is reduced in higher ionic strength. Our results indicate that the structure of PDC may not always maintain its ~ 10 MDa organization, but is rather variable. We propose that the flexible nature of PDC may allow modulation of its activity.

scholarly journals Regulation of pyruvate dehydrogenase kinase activity from pig kidney cortex

1992 ◽  
Vol 288 (2) ◽  
pp. 369-373 ◽  
Author(s):  
T Pawelczyk ◽  
M S Olson
Keyword(s):  
Ionic Strength ◽  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Kidney Cortex ◽  
Small Range ◽  
High Concentration ◽  
Optimum Activity ◽  
Pig Kidney

The activity of pyruvate dehydrogenase (PDH) kinase in the purified PDH complex from pig kidney is sensitive to changes in ionic strength. The enzyme has optimum activity within a small range of ionic strength (0.03-0.05 M). An increase in ionic strength from 0.04 M to 0.2 M lowers the activity of PDH kinase by 32% and decreases the Km for ATP from 25 microM to 10 microM. At constant ionic strength (0.15 M) the enzyme has optimum activity over a broad pH range (7.2-8.0). The PDH kinase is stimulated 2.2-fold by 20 mM-K+, whereas Na+ even at high concentration (80 mM) has no effect on the enzyme activity. The stimulation of PDH kinase by K+ is not dependent on pH and ionic strength. PDH kinase is inhibited by HPO4(2-) in the presence of K+, whereas HPO4(2-) has no effect on the activity of this enzyme in the absence of K+. HPO4(2-) at concentrations of 2 and 10 mM inhibits PDH kinase by 28% and 55% respectively. The magnitude of this inhibition is not dependent on the ATP/ADP ratio. Inhibition by HPO4(2-) in the concentration range 0-10 mM is non-competitive with respect to ATP, and becomes mixed-type at concentrations over 10 mM. The Ki for HPO4(2-) is 10 mM. When HPO4(2-) is replaced by SO4(2-), the same effects on the activity of PDH kinase are observed. PDH kinase is also inhibited by Cl-. In the presence of 80 mM-Cl- the PDH kinase is inhibited by 40%. The inhibition by Cl- is not dependent on K+. In conclusion, we postulate that changes in phosphate concentrations may play a significant role in the regulation of PDH kinase activity in vivo.

scholarly journals The plasticity of the pyruvate dehydrogenase complex confers a labile structure that is associated with its catalytic activity

PLoS ONE ◽  
2020 ◽  
Vol 15 (12) ◽  
pp. e0243489
Author(s):  
Jaehyoun Lee ◽  
Seunghee Oh ◽  
Saikat Bhattacharya ◽  
Ying Zhang ◽  
Laurence Florens ◽  
...  
Keyword(s):  
Ionic Strength ◽  
Pyruvate Dehydrogenase ◽  
Biochemical Analysis ◽  
Pyruvate Dehydrogenase Complex ◽  
Size Exclusion ◽  
Dependent Manner ◽  
Acetyl Coa ◽  
Cell Extracts ◽  
Exclusion Chromatography ◽  
Dehydrogenase Complex

The pyruvate dehydrogenase complex (PDC) is a multienzyme complex that plays a key role in energy metabolism by converting pyruvate to acetyl-CoA. An increase of nuclear PDC has been shown to be correlated with an increase of histone acetylation that requires acetyl-CoA. PDC has been reported to form a ~ 10 MDa macromolecular machine that is proficient in performing sequential catalytic reactions via its three components. In this study, we show that the PDC displays size versatility in an ionic strength-dependent manner using size exclusion chromatography of yeast cell extracts. Biochemical analysis in combination with mass spectrometry indicates that yeast PDC (yPDC) is a salt-labile complex that dissociates into sub-megadalton individual components even under physiological ionic strength. Interestingly, we find that each oligomeric component of yPDC displays a larger size than previously believed. In addition, we show that the mammalian PDC also displays this uncommon characteristic of salt-lability, although it has a somewhat different profile compared to yeast. We show that the activity of yPDC is reduced in higher ionic strength. Our results indicate that the structure of PDC may not always maintain its ~ 10 MDa organization, but is rather variable. We propose that the flexible nature of PDC may allow modulation of its activity.

scholarly journals Kinetic properties of the partially purified pyruvate dehydrogenase complex of ox brain

1973 ◽  
Vol 131 (1) ◽  
pp. 31-37 ◽  
Author(s):  
John P. Blass ◽  
Carole A. Lewis
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Tricarboxylic Acid Cycle ◽  
Pyruvate Dehydrogenase Complex ◽  
Kinetic Properties ◽  
Acetyl Coa ◽  
Ph Optimum ◽  
Michaelis Constants ◽  
High Concentrations ◽  
Similar Preparation ◽  
Dehydrogenase Complex

The properties of a purified preparation of the pyruvate dehydrogenase complex from ox brain have been compared with those of a similar preparation from ox kidney. A broad pH optimum around 7.8, similar dependence on ionic strength, and independence of the nature of the buffer anions or cations characterized preparations from both tissues. Michaelis constants for the binding of pyruvate, thiamin pyrophosphate, NAD+ and CoA were also similar. Enzyme from both tissues was inhibited by NADH, by copper and other heavy metals, by high concentrations of tricarboxylic acid-cycle intermediates, and by preincubation with ATP. Acetyl-CoA itself did not appear to inhibit these preparations, although some commercial preparations of acetyl-CoA did contain an inhibitor. Although oxaloacetate and α-oxobutyrate were weak inhibitors, a number of other α-oxo acids including phenylpyruvate did not inhibit. The properties of the pyruvate dehydrogenase complex from brain and kidney appeared similar.

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