Geranylgeranyl pyrophosphate synthetase lacking geranyl-transferring activity from Micrococcus luteus.
Geranyl pyrophosphate synthetase, which catalyzes the condensation of isopentenyl pyrophosphate with dimethylallyl pyrophosphate to give geranyl pyrophosphate, was purified 490-fold from Micrococcus luteus extracts by DEAE-Sephadex, hydroxylapatite, and Sephadex G-100 column chromatography. The enzyme has a pH optimum at 7.7 and the molecular weight was estimated to be 70,000 by Sephadex gel filtration. The Km values for isopentenyl pyrophosphate and dimethylallyl pyrophosphate were 8 microM and 62 microM, respectively. The enzyme required Mg2+ for maximum activity. Tween 80 showed a stimulative effect whereas Triton X-100 was rather inhibitory on the enzyme activity. Inorganic pyrophosphate and iodoacetamide were both potent inhibitors of the enzyme. The purified enzyme fraction was also capable of catalyzing the synthesis of geranylgeranyl pyrophosphate from isopentenyl pyrophosphate and farnesyl pyrophosphate, but lacked geranyl-transferring activity. The catalytic activities of geranylgeranyl pyrophosphate synthesis and geranyl pyrophosphate synthesis were affected differently by iodoacetamide and Triton X-100. This enzyme fraction may be a mixture of two enzymes, geranyl pyrophosphate synthetase and geranylgeranyl pyrophosphate synthetase catalyzing the reactions of C5 greater than C10 and C15 greater than C20, respectively, or a single enzyme with two independent catalytic sites responsible for the C5 greater than C10 and C15 greater than C20 reactions. In any case, the existence of a new geranylgeranyl pyrophosphate synthetase different from the known geranylgeranyl pyrophosphate synthetase catalyzing the continuous condensation reaction of C5 greater than C10 greater than C15 greater than C20 was demonstrated.
D019883:Alkyl and Aryl Transferases; D004122:Dimethylallyltranstransferase / Q000378:metabolism; D051228:Geranyltranstransferase; D007460:Iodoacetamide / Q000494:pharmacology; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008837:Micrococcus / Q000201:enzymology*; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D011106:Polyisoprenyl Phosphates / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism; D013379:Substrate Specificity; D014166:Transferases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*
D011106:Polyisoprenyl Phosphates; D011092:Polyethylene Glycols; D017830:Octoxynol; D014166:Transferases; D019883:Alkyl and Aryl Transferases; D004122:Dimethylallyltranstransferase; D051228:Geranyltranstransferase; D008274:Magnesium; C002963:geranylgeranyl pyrophosphate; D007460:Iodoacetamide