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Propionyl-CoA condensing enzyme from Ascaris muscle mitochondria. II. Coenzyme A modulation.
- Source :
-
Archives of biochemistry and biophysics [Arch Biochem Biophys] 1991 Feb 15; Vol. 285 (1), pp. 166-71. - Publication Year :
- 1991
-
Abstract
- The propionyl-CoA condensing enzyme which catalyzes the first step in the biosynthesis of 2-methylbutyrate and 2-methylvalerate by Ascaris muscle appears to exist in at least three forms in the mitochondria of this parasitic nematode. Two forms, A and B, were separated by ion exchange chromatography on CM-Sephadex. Chromatography on phosphocellulose resulted in the recovery of one minor peak (I) and two major peaks with activity (II and III). A and B as well as I, II, and III differed in their specific activities. Forms B and III were the most retained by their resins, and were the most active forms of the enzyme in each case. Inhibition studies with metabolites from Ascaris mitochondria indicate that CoASH, a product of the condensation reaction, and acetyl-CoA are effective inhibitors of the condensing and thiolytic activities of the Ascaris enzyme, respectively. Incubation of the active enzyme form B for 2 h with 0.1 mM CoASH at room temperature under nitrogen caused the loss of 92% of the propionyl-CoA condensing activity and 67% of the thiolase activity when assayed in standard mixtures. The propionyl-CoA condensing enzyme exhibited a hyperbolic dependence of the condensation velocity to changes in substrate concentration. However, in the presence of CoASH the Michaelis-Menten kinetics was transformed into a sigmoidal kinetics indicating a deviation from a simple product inhibition. Inactivation of the most active forms of the enzyme with CoASH was accompanied by (a) a change in the chemical reactivity of the protein toward p-chloromurcuribenzoate, (b) a change in the uv-visible spectrum of the protein, and (c) a change in the elution patterns from both CM-Sephadex and phosphocellulose column chromatography, where-upon one, two, or more protein peaks were obtained. The several protein peaks resolved by rechromatography of the [14C]CoASH-inactivated enzyme III on phosphocellulose had different CoASH contents. The elution positions were correlated with the less active forms (I and II) having increased [14C]CoASH activities. Similarly, the two peaks isolated upon rechromatography of the CoASH-partially inactivated enzyme B on CM-Sephadex had different isotope contents and the elution position of enzyme A corresponded to the less active form. The results described indicate that the CoASH modification of Ascaris propionyl-CoA condensing enzyme may be responsible for the existence of several forms of the enzyme and might represent a mode of control by chemically modulating the amount of the active forms of the enzyme.
- Subjects :
- Acetyl-CoA C-Acetyltransferase metabolism
Animals
Ascaris drug effects
Coenzyme A pharmacology
Enzyme Activation drug effects
Isoenzymes metabolism
Kinetics
Mitochondria, Muscle drug effects
Nitrogen pharmacology
Phencyclidine analogs & derivatives
Phencyclidine pharmacology
Alcohol Oxidoreductases metabolism
Ascaris enzymology
Mitochondria, Muscle enzymology
Subjects
Details
- Language :
- English
- ISSN :
- 0003-9861
- Volume :
- 285
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Archives of biochemistry and biophysics
- Publication Type :
- Academic Journal
- Accession number :
- 1671327
- Full Text :
- https://doi.org/10.1016/0003-9861(91)90345-j