Tritium isotope effects in the measurement of the glycerol phosphate and dihydroxyacetone phosphate pathways of glycerolipid biosynthesis in rat liver

Manning, Ray; Brindley, David N.

doi:10.1042/bj1301003

Cited by 86 publications

(31 citation statements)

References 25 publications

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“…We also found decreased circulating TG levels in fl d mice ( Table 2 ), which could affect the availability of substrate for cardiac work. We only measured serum FA levels after 2 h of food deprivation in the the specifi c radioactivity in the glycerol 3-phosphate precursor pool is also measured ( 71 ). This specifi c activity depends on the relative activities of glycerol kinase in the fl d and control hearts and on the rate of substrate cycling between glycerol 3-phosphate and dihydroxyacetone phosphate.…”

Section: Examination Of the Downstream Signaling Effects Of Aberrant mentioning

confidence: 99%

Relationship of glucose and oleate metabolism to cardiac function in lipin-1 deficient (fld) mice

Kok

Kienesberger

Dyck

et al. 2012

Journal of Lipid Research

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of cardiac ATP production ( 1 ). FAs are also incorporated into triacylglycerol (TG), which is stored in lipid droplets. Cardiac TG turnover is rapid ( 2 ) and FAs released from TG can contribute up to 10-20% of ATP production ( 3 ). Plasma FA and TG levels rise in insulin resistance and diabetes ( 4, 5 ), and cardiac FA utilization can increase to supply as much as 90% of ATP ( 6, 7 ). Despite this increase in FAO, FA uptake can exceed the ability of the heart to utilize FAs, leading to excess accumulation of TG in cardiomyocytes ( 6-9 ). The compensatory changes that occur in response to excessive FA availability eventually become maladaptive, and the heart is locked in a pathological and infl exible metabolic state that contributes to cardiac dysfunction ( 6, 7, 10 ). As such, maintenance of metabolic fl exibility is essential for normal cardiac function. Whereas many proteins have been identifi ed to contribute to cardiac FA uptake and metabolism ( 1, 10 ), relatively little information is available about the role of lipin-1 in the heart. Lipin-1 is an important protein involved in regulating both TG synthesis and FAO in other organs ( 11 ).Instead of being considered as mutually antagonistic processes, FAO and TG synthesis are now widely viewed as being companion pathways because augmenting cardiac TG synthesis increases FAO ( 12 ). Moreover, FAs released through lipolysis of endogenous TG stores contribute signifi cantly to FAO ( 3,13 ). Lipin-1 could play a prominent role in this regulation as it is a bifunctional protein involved in regulating both TG synthesis and FAO in the The constant requirement for ATP in the beating heart is satisfi ed by the utilization of diverse substrates, such as fatty acids (FA), glucose, lactate, ketones, and amino acids, with FA oxidation (FAO) providing the majority (50-75%)

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Section: Examination Of the Downstream Signaling Effects Of Aberrant mentioning

confidence: 99%

Relationship of glucose and oleate metabolism to cardiac function in lipin-1 deficient (fld) mice

Kok

Kienesberger

Dyck

et al. 2012

Journal of Lipid Research

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“…The radioactivity in a sample of the final supernatant was measured by the method of Manning & Brindley (1972). Preliminary experiments, designed to measure the loss of bicarbonate during an incubation, involved incubation of the assay medium without enzyme for 30min at 37°C.…”

Section: Assay Of Acetyl-coa Car4oxylasementioning

confidence: 99%

Purification and some properties of acetyl-coenzyme A carboxylase from rabbit mammary gland

Manning¹,

Dils²,

Mayer³

1976

Biochemical Journal

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1. Acetyl-Coa carboxylase from lactating-rabbit mammary gland was purified to homogeneity by the criterion of polyacrylamide-gel electrophoresis in the presence of sodium dodecyl sulphate. 2. Use of phosphate buffer throughout the purification gave low recovery of enzyme. Consequently, Tris buffers were used in the extraction and in selected stages of the purification procedure. 3. The purified enzyme had a specific activity of 5.15 +/- 0.3 μmol of bicarbonate incorporated/min per mg of protein (mean +/- S.E.M. of five preparations). This represents a purification of 257 +/- 16-fold and a yield of 4.3 +/- 0.13%. 4. The kinetic parameters of the purified enzyme were similar to those reported for the enzyme from other tissue sources. 5. The enzyme was assayed by a spectrophotometric assay and by a [14C]bicarbonate-fixation assay. Short incubation were used in the radio-chemical assay to avoid substantial loss of [14C]bicarbonate.

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“…The question of the relative contributions of the GP and acyl DHAP pathways to phosphoglycerolipid and triacylglycerol biosynthesis in eukaryotes remains an important one (6,29,30,33). In …”

Section: Resultsmentioning

confidence: 99%

“…The enzymatic conversion of alkyl DHAP to glycerol ether lipids proceeds via reduction of alkyl DHAP to 1-alkyl GP followed by acylation (13). The importance of the acyl DHAP pathway for the biosynthesis of non-ether glycerolipids is controversial (6,29,30,33,34,35). Except for some anaerobic bacteria which contain plasmalogens but not alkyl glycerol ethers, both aerobic bacteria and facultative anaerobes, as well as plants, lack ether lipids (9, 28).…”

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The acyl dihydroxyacetone phosphate pathway enzymes for glycerolipid biosynthesis are present in the yeast Saccharomyces cerevisiae

et al. 1992

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The presence of the acyl dihydroxyacetone phosphate (acyl DHIAP) pathway in yeasts was investigated by examining three key enzyme activities of this pathway in Saccharomyces cerevisiae. In the total membrane fraction of S. cerevisiae, we confirmed the presence of both DHAP acyltransferase (DHAPAT; Km = 1.27 mM; Vmax = 5.9 nmol/min/mg of protein) and sn-glycerol 3-phosphate acyltransferase (GPAT; Km = 0.28 mM; Vmax = 12.6 nmol/min/mg of protein). The properties of these two acyltransferases are similar with respect to thermal stability and optimum temperature of activity but differ with respect to pH optimum (6.5 for GPAT and 7.4 for DHAPAT) and sensitivity toward the sulfhydryl blocking agent N-ethylmaleimide. Total membrane fraction of S. cerevisiae also exhibited acyl/alkyl DHAP reductase (EC 1.1.1.101) activity, which has not been reported previously. The reductase has a Vm, of 3.8 nmol/min/mg of protein for the reduction of hexadecyl DHLAP (Km = 15 ,uM) by NADPH (Km = 20 ,uM). Both acyl DHAP and alkyl DHAP acted as substrates. NADPH was the specific cofactor. Divalent cations and N-ethylmaleimide inhibited the enzymatic reaction.Reductase activity in the total membrane fraction from aerobically grown yeast cells was twice that from anaerobically grown cells. Similarly, DHAPAT and GPAT activities were also greater in aerobically grown yeast cells. The presence of these enzymes, together with the absence of both ether glycerolipids and the ether lipid-synthesizing enzyme (alkyl DHIAP synthase) in S. cerevisiae, indicates that non-ether glycerolipids are synthesized in this organism via the acyl DHAP pathway.Biosynthesis of phosphatidic acid, the precursor of glycerolipids in all organisms, proceeds via two different routes. In the glycerol phosphate pathway, phosphatidic acid is biosynthesized via the stepwise enzymatic acylation of snglycerol 3-phosphate (GP) by long-chain fatty acyl coenzyme A's (acyl-CoAs). In animals, phosphatidate is also biosynthesized by an alternate pathway, the acyl dihydroxyacetone phosphate (acyl DHAP) pathway. In this pathway, DHAP is acylated by a long-chain fatty acyl-CoA to form acyl DHAP. This compound is then reduced to 1-acyl GP, which is further acylated to yield phosphatidate (Fig. 1). The GP pathway is universal to all organisms, including yeasts (19,27). In contrast, the acyl DHAP pathway is absent in bacteria and plants (11, 41). In animals, the acyl DHAP pathway is obligatory for ether lipid biosynthesis; the ether bond is synthesized by the substitution of a long-chain alcohol for the acyl group of acyl DHAP, with the release of free fatty acid to form alkyl DHAP (13). The enzymatic conversion of alkyl DHAP to glycerol ether lipids proceeds via reduction of alkyl DHAP to 1-alkyl GP followed by acylation (13). The importance of the acyl DHAP pathway for the biosynthesis of non-ether glycerolipids is controversial (6,29,30,33,34,35). Except for some anaerobic bacteria which contain plasmalogens but not alkyl glycerol ethers, both aerobic bacteria and facultative anaerobe...

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Tritium isotope effects in the measurement of the glycerol phosphate and dihydroxyacetone phosphate pathways of glycerolipid biosynthesis in rat liver

Cited by 86 publications

References 25 publications

Relationship of glucose and oleate metabolism to cardiac function in lipin-1 deficient (fld) mice

Relationship of glucose and oleate metabolism to cardiac function in lipin-1 deficient (fld) mice

Purification and some properties of acetyl-coenzyme A carboxylase from rabbit mammary gland

The acyl dihydroxyacetone phosphate pathway enzymes for glycerolipid biosynthesis are present in the yeast Saccharomyces cerevisiae

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