Abstract
Long chain cis-prenyltransferase in rat liver microsomes was studied using various allylic isoprenoid substrates. Microsomes could utilize trans-geranyl pyrophosphate, but not cis-geranyl pyrophosphate for polyprenyl pyrophosphate synthesis. Both trans, trans-farnesyl pyrophosphate and trans,cis-farnesyl pyrophosphate were used as substrates with Km values of 24 and 5 microM, respectively. trans,trans,cis-Geranylgeranyl pyrophosphate could be used as substrate with an apparent Km of 36 microM. trans,trans,trans-Geranylgeranyl pyrophosphate was also utilized as substrate, but with a very low affinity. After pulse labeling for 4 min, using [3H]isopentenyl pyrophosphate and trans,trans-farnesyl pyrophosphate, the only product formed was trans,trans,cis-geranylgeranyl pyrophosphate, which, upon chasing, yielded polyprenyl pyrophosphate. Independent of the nature of the substrate used, even in the case of polyprenyl 12-pyrophosphate and all-trans-nonaprenyl pyrophosphate, the chain lengths of the products were identical, i.e. polyprenyl pyrophosphates with 15-18 isoprene residues. Microsomes were able to synthesize trans,trans-farnesyl pyrophosphate using trans-geranyl pyrophosphate as substrate. The results indicate that rat liver microsomes contain a farnesyl pyrophosphate synthase activity and that the reaction catalyzed by cis-prenyltransferase may consist of two individual steps, i.e. synthesis of trans,trans,cis-geranylgeranyl pyrophosphate and elongation of this product to long chain polyprenyl pyrophosphates.
Highlights
Somes was studied usingvariousallylic isoprenoid cis-Prenyltransferases havebeen isolatedand characterized substrates
Trans-farnesyl pyrophosphate and trans,cis-farnesyl pyrophosphate were used as substrates with K,values of 24 and 5 p M, respectively. trans,trans,cis-Geranylgeranyl pyrophosphate could be used as substrate with tions from hen oviduct, rat liver, rat brain, Ehrlich ascites tumor cells, and spermatogeniccells, certain propertiesof the enzyme activity havebeen analyzed using FPP and GGPP as substrates [7,8,9,10,11,12]
The results indicate that rat liver micro- mediate in the biosynthesis of long chain polyprenyl pyrosomes contain a farnesyl pyrophosphate synthase ac- phosphates
Summary
All unlabeledpyrophosphate substrates were synthesized from the corresponding alcohols was extended to 120 min, no product eluting early could be observed, and all the radioactivity was associatedwith the long chain polyprenols (Fig. 1C). These results indicate that the product after a short incubation is GGPP, which can be metabolized to long chain products containing isoprene residues. 3. Substrate specificityof rat liver cis-prenyltransferase.Microsomes were incubated with increasing amounts of various allylic substrates and saturating amounts of [3H]IPP.The amount of productformed (determined asthe total extractable radioactivity) was plotted against substrate concentration.
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