Glyoxysomal acetoacetyl-CoA thiolase and 3-oxoacyl-CoA thiolase from sunflower cotyledons.
Following chromatography on hydroxyapatite, the elution profile of the thiolase activity of the glyoxysomal fraction from sunflower (Helianthus annuus L.) cotyledons exhibited two peaks when the enzyme activity was assayed with acetoacetyl-CoA as substrate. Only one of these two activity peaks was detectable when a long-chain thiolase substrate was used in the activity assay. The proteins (thiolase I and thiolase II) underlying the two activity peaks detected with acetoacetyl-CoA were of glyoxysomal origin. They were purified using glyoxysomal matrices as starting material, and biochemically characterized. Thiolase I is an acetoacetyl-CoA thiolase (EC 2.3.1.9) exhibiting activity only towards acetoacetyl-CoA (Km = 11 microM). Its contribution to the total glyoxysomal thiolytic activity towards acetoacetyl-CoA amounted to about 15%. Thiolase II is a 3-oxoacyl-CoA thiolase (EC 2.3.1.16). The activity of the enzyme towards 3-oxoacyl-CoAs increased with increasing chain length of the substrate. Thiolase II exhibited a Km value of 27 microM with acetoacetyl-CoA as substrate. and Km values between 3 and 7 microM with substrates having a carbon chain length from 6 to 16 carbon atoms. The thiolase activity of the glyoxysomes towards acetoacetyl-CoA and 3-oxopalmitoyl-CoA exceeded the glyoxysomal butyryl-CoA and palmitoyl-CoA beta-oxidation rates, respectively, by about 10-fold at all substrate concentrations employed (1-15 microM).
Oeljeklaus
Silke
S
Institut für Botanik, Universität Münster, Germany.
Fischer
Kerstin
K
Gerhardt
Bernt
B
D000101:Acetyl-CoA C-Acetyltransferase / Q000302:isolation & purification / Q000378:metabolism*; D000102:Acetyl-CoA C-Acyltransferase / Q000302:isolation & purification / Q000378:metabolism*; D000214:Acyl Coenzyme A / Q000378:metabolism; D018548:Cotyledon / Q000201:enzymology*; D005227:Fatty Acids / Q000378:metabolism; D020676:Glyoxysomes / Q000201:enzymology*; D006368:Helianthus / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D010171:Palmitoyl Coenzyme A / Q000378:metabolism
D000214:Acyl Coenzyme A; D005227:Fatty Acids; C010667:acetoacetyl CoA; D010171:Palmitoyl Coenzyme A; C024343:butyryl-coenzyme A; D000102:Acetyl-CoA C-Acyltransferase; D000101:Acetyl-CoA C-Acetyltransferase