Determination of isopentenyl diphosphate and farnesyl diphosphate in tissue samples with a comment on secondary regulation of polyisoprenoid biosynthesis
A double-isotope dilution procedure is described for the determination of two isoprenoid precursors, isopentenyl and farnesyl diphosphate. Recovery of each is determined by the addition of the appropriate radioactive diphosphate to the tissue sample. After partial purification, each is coupled by a prenyltransferase with a cosubstrate of known specific activity. The products, doubly labeled farnesyl and geranylgeranyl diphosphates, are cleaved to the parent alcohols by alkaline phosphatase. The resulting polyprenols are isolated by reversed-phase thin-layer chromatography and their radioisotopic content is determined. The levels of these precursors have been measured in livers of rats and mice that have been maintained on several different diets. The concentration of each was about 0.5 mumol/g wet tissue and varied as much as 10-fold under the different test conditions. The levels of isopentenyl diphosphate isomerase, farnesyl diphosphate synthetase, and squalene synthetase were also measured in these animals. The changes in levels of these enzymes, in conjunction with the variation in substrate concentrations, are such that they could substantially influence the rate of cholesterol synthesis in liver.
- Research Organization:
- Univ. of Utah School of Medicine, Salt Lake City (USA)
- OSTI ID:
- 6605725
- Journal Information:
- Anal. Biochem.; (United States), Vol. 173:2
- Country of Publication:
- United States
- Language:
- English
Similar Records
Isopentenyl diphosphate (IPP)-bypass mevalonate pathways for isopentenol production
Structural and thermodynamic basis of the inhibition of Leishmania major farnesyl diphosphate synthase by nitrogen-containing bisphosphonates
Related Subjects
CHOLESTEROL
BIOSYNTHESIS
ORGANIC PHOSPHORUS COMPOUNDS
FRACTIONATION
ALKALINE PHOSPHATASE
ISOMERASES
ISOTOPE DILUTION
LIGASES
LIVER
MICE
PRECURSOR
RATS
SCINTILLATION COUNTING
THIN-LAYER CHROMATOGRAPHY
TRANSFERASES
ANIMALS
BODY
CHROMATOGRAPHY
COUNTING TECHNIQUES
DIGESTIVE SYSTEM
ENZYMES
ESTERASES
GLANDS
HYDROLASES
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
MAMMALS
ORGANIC COMPOUNDS
ORGANS
PHOSPHATASES
RODENTS
SEPARATION PROCESSES
STEROIDS
STEROLS
SYNTHESIS
TRACER TECHNIQUES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques