Plant acyl-CoA oxidase. Purification, characterization, and monomeric apoprotein.
Purified glyoxysomes from cotyledons of germinating cucumber seedlings were used as a source to separate matrix enzymes of the organelle by hydrophobic chromatography. Glyoxysomal acyl-CoA oxidase eluted from the column like hydrophobic proteins and exhibited an Mr of 150,000. An oxidase with identical properties could be prepared in large quantities by a purification procedure starting with crude extracts from cotyledons of 4-day-old etiolated seedlings. The purification procedure included chromatography on phenyl-Sepharose and hydroxylapatite and molecular sieving. 1500-fold purification led to an enzyme of apparent homogeneity characterized by a specific activity of 27 units/mg of protein. Plant acyl-CoA oxidase is a homodimer with a subunit of Mr 72,000. Monospecific antibodies raised in rabbits were used to reveal dissimilarity to the fungal oxidase. The plant enzyme also differed markedly in molecular structure and amino acid composition from the liver peroxisomal enzyme. Glyoxysomal acyl-CoA oxidase acts selectively on fatty acyl-CoAs with 16 or 18 C atoms, cis-9-unsaturated esters with a C16 or C18 acyl moiety being converted with higher rates than saturated or polyunsaturated fatty acyl-CoAs. Besides the enzymatically active organellar form of acyl-CoA oxidase, the monomeric apoprotein was detected when short-term labeling of cotyledons in vivo was performed. The apoprotein (immunoprecipitable by antibodies raised against the glyoxysomal enzyme) did not differ in size from the subunit of the glyoxysomal dimeric enzyme.
D000214:Acyl Coenzyme A / Q000378:metabolism; D042962:Acyl-CoA Oxidase; D000596:Amino Acids / Q000032:analysis; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005779:Immunodiffusion; D007526:Isoelectric Point; D046911:Macromolecular Substances; D008970:Molecular Weight; D010088:Oxidoreductases / Q000302:isolation & purification*; D010944:Plants / Q000201:enzymology*; D013379:Substrate Specificity
D000214:Acyl Coenzyme A; D000596:Amino Acids; D046911:Macromolecular Substances; D010088:Oxidoreductases; D042962:Acyl-CoA Oxidase