Isopentenyl pyrophosphate isomerase and prenyltransferase from tomato fruit plastids.
Isopentenyl pyrophosphate isomerase has been isolated from an extract of tomato fruit plastids and purified 245-fold by fractionation with ammonium sulfate, gel filtration on Bio-Gel A 1.5m, ion-exchange chromatography on DEAE-cellulose, gel filtration on Sephadex G-100, and chromatofocusing. Gel filtration on Sephadex G-100 separated the isopentenyl pyrophosphate isomerase from a prenyltransferase fraction that catalyzed the conversion of isopentenyl pyrophosphate to acid-labile compounds in the presence of dimethylallyl, geranyl, or farnesyl pyrophosphates. The molecular weights of the isopentenyl pyrophosphate isomerase and prenyltransferase were determined to be 34,000 and 64,000, respectively, by gel filtration on Sephadex G-100. The only cofactor required by either the isomerase or the prenyltransferase was a divalent cation, either Mg2+ or Mn2+. Isopentenyl pyrophosphate isomerase could also be totally inactivated by 1 X 10(-3) M iodoacetamide, and this property was utilized in the assay of prenyltransferase activity in the presence of contaminating isomerase. The inactivation of isomerase by iodoacetamide is consistent with the stabilization of isopentenyl pyrophosphate isomerase by dithiothreitol. The Km of isopentenyl pyrophosphate isomerase for isopentenyl pyrophosphate was found to be 5.7 X 10(-6).
D019748:Carbon-Carbon Double Bond Isomerases; D002736:Chloroplasts / Q000201:enzymology; D002845:Chromatography / Q000379:methods; D003067:Coenzymes / Q000378:metabolism; D004122:Dimethylallyltranstransferase / Q000302:isolation & purification*; D011756:Diphosphates / Q000494:pharmacology; D045782:Hemiterpenes; D007460:Iodoacetamide / Q000494:pharmacology; D007525:Isoelectric Focusing; D007535:Isomerases / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D007700:Kinetics; D008970:Molecular Weight; D010944:Plants / Q000201:enzymology*; D014166:Transferases / Q000302:isolation & purification*
D003067:Coenzymes; D011756:Diphosphates; D045782:Hemiterpenes; D014166:Transferases; D004122:Dimethylallyltranstransferase; D007535:Isomerases; D019748:Carbon-Carbon Double Bond Isomerases; C021453:isopentenyldiphosphate delta-isomerase; D007460:Iodoacetamide