Fatty acid metabolism in sn-glycerol-3-phosphate acyltransferase (plsB) mutants.

Fatty acid metabolism was examined in Escherichia coli plsB mutants that were conditionally defective in sn-glycerol-3-phosphate acyltransferase activity. The fatty acids synthesized when acyl transfer to glycerol-3-phosphate was inhibited were preferentially transferred to phosphatidylglycerol. A comparison of the ratio of phospholipid species labeled with 32Pi and [3H]acetate in the presence and absence of glycerol-3-phosphate indicated that [3H]acetate incorporation into phosphatidylglycerol was due to fatty acid turnover. A significant contraction of the acetyl coenzyme A pool after glycerol-3-phosphate starvation of the plsB mutant precluded the quantitative assessment of the rate of phosphatidylglycerol fatty acid labeling. Fatty acid chain length in membrane phospholipids increased as the concentration of the glycerol-3-phosphate growth supplement decreased, and after the abrupt cessation of phospholipid biosynthesis abnormally long chain fatty acids were excreted into the growth medium. These data suggest that the acyl moieties of phosphatidylglycerol are metabolically active, and that competition between fatty acid elongation and acyl transfer is an important determinant of the acyl chain length in membrane phospholipids.     

A missense mutation accounts for the defect in the glycerol-3-phosphate acyltransferase expressed in the plsB26 mutant

The sn-glycerol-3-phosphate acyltransferase (plsB) catalyzes the first step in membrane phospholipid formation. A conditional Escherichia coli mutant (plsB26) has a single missense mutation (G1045A) predicting the expression of an acyltransferase with an Ala349Thr substitution. The PlsB26 protein had a significantly reduced glycerol-3-phosphate acyltransferase specific activity coupled with an elevated Km for glycerol-3-phosphate.     

sn-Glycerol-3-phosphate auxotrophy of plsB strains of Escherichia coli: evidence that a second mutation, plsX, is required.

sn-Glycerol-3-phosphate auxotrophs defective in phospholipid synthesis contain a Km-defective sn-glycerol-3-phosphate acyltransferase. Detailed genetic analysis revealed that two mutations were required for the auxotrophic phenotype. One mutation, in the previously described plsB locus (sn-glycerol-3-phosphate acyltransferase structural gene), mapped near min 92 on the Escherichia coli linkage map. Isolation of Tn10 insertions cotransducible with the auxotrophy in phage P1 crosses revealed that a second mutation was required with plsB26 to confer the sn-glycerol-3-phosphate auxotrophic phenotype. This second locus, plsX, mapped between pyrC and purB near min 24 on the E. coli linkage map. Tn10 insertions near plsX allowed detailed mapping of the genetic loci in this region. A clockwise gene order putA pyrC flbA flaL flaT plsX fabD ptsG thiK purB was inferred from results of two- and three-factor crosses. Strains harboring the four possible configurations of the mutant and wild-type plsB and plsX loci were constructed. Isogenic plsB+ plsX+, plsB+ plsX50, and plsB26 plsX+ strains grew equally well on glucose minimal medium without sn-glycerol-3-phosphate. In addition, plsX or plsX+ had no apparent effect on sn-glycerol-3-phosphate acyltransferase activity measured in membrane preparations. The molecular basis for the plsX requirement for conferral of sn-glycerol-3-phosphate auxotrophy in these strains remains to be established.     

Glycerol-3-phosphatase of Corynebacterium glutamicum

Formation of glycerol as by-product of amino acid production by Corynebacterium glutamicum has been observed under certain conditions, but the enzyme(s) involved in its synthesis from glycerol-3-phosphate were not known. It was shown here that cg1700 encodes an enzyme active as a glycerol-3-phosphatase (GPP) hydrolyzing glycerol-3-phosphate to inorganic phosphate and glycerol. GPP was found to be active as a homodimer. The enzyme preferred conditions of neutral pH and requires Mg2? or Mn2? for its activity. GPP dephosphorylated both L- and D-glycerol-3-phosphate with a preference for the D-enantiomer. The maximal activity of GPP was estimated to be 31.1 and 1.7 U mg?1 with K(M) values of 3.8 and 2.9 mM for DL- and L-glycerol-3-phosphate, respectively. For physiological analysis a gpp deletion mutant was constructed and shown to lack the ability to produce detectable glycerol concentrations. Vice versa, gpp overexpression increased glycerol accumulation during growth in fructose minimal medium. It has been demonstrated previously that intracellular accumulation of glycerol-3-phosphate is growth inhibitory as shown for a recombinant C. glutamicum strain overproducing glycerokinase and glycerol facilitator genes from E. coli in media containing glycerol. In this strain, overexpression of gpp restored growth in the presence of glycerol as intracellular glycerol-3-phosphate concentrations were reduced to wild-type levels. In C. glutamicum wild type, GPP was shown to be involved in utilization of DL-glycerol-3-phosphate as source of phosphorus, since growth with DL-glycerol-3-phosphate as sole phosphorus source was reduced in the gpp deletion strain whereas it was accelerated upon gpp overexpression. As GPP homologues were found to be encoded in the genomes of many other bacteria, the gpp homologues of Escherichia coli (b2293) and Bacillus subtilis (BSU09240, BSU34970) as well as gpp1 from the plant Arabidosis thaliana were overexpressed in E. coli MG1655 and shown to significantly increase GPP activity.     

A gene (plsD) from Clostridium butyricum that functionally substitutes for the sn-glycerol-3-phosphate acyltransferase gene (plsB) of Escherichia coli.

The sn-glycerol-3-phosphate acyltransferase (plsB) of Escherichia coli is a key regulatory enzyme that catalyzes the first committed step in phospholipid biosynthesis. We report the initial characterization of a novel gene (termed plsD) from Clostridium butyricum, cloned based on its ability to complement the sn-glycerol-3-phosphate auxotrophic phenotype of a plsB mutant strain of E. coli. Unlike the 83-kDa PlsB acyltransferase from E. coli, the predicted plsD open reading frame encoded a protein of 26.5 kDa. Two regions of strong homology to other lipid acyltransferases, including PlsB and PlsC analogs from mammals, plants, yeast, and bacteria, were identified. PlsD was most closely related to the 1-acyl-sn-glycerol-3-phosphate acyltransferase (plsC) gene family but did not complement the growth of plsC(Ts) mutants. An in vivo metabolic labeling experiment using a plsB plsX plsC(Ts) strain of E. coli confirmed that the plsD expression restored the ability of the cells to synthesize 1-acyl-glycerol-3-phosphate. However, glycerol-3-phosphate acyltransferase activity was not detected in vitro in assays using either acyl-acyl carrier protein or acyl coenzyme A as the substrate.     

Uptake and dissimilation of glycerol by wild type and glycerol nonutilizing strains of Neurospora crassa

Summary Seven mutant strains defective for utilization of glycerol, glyceraldehyde or dihydroxyacetone were isolated. One strain was deficient for NAD-linked glycerol-3-phosphate dehydrogenase, two for glycerol kinase, and four had no detected enzymatic deficiency, although one of the latter strains was deficient in glycerol uptake. Glycerol uptake was increased by incubation in glycerol, glycerol-3-phosphate, erythritol, and propanediol, and was protein-mediated below 0.14 mM glycerol, but at higher concentrations free diffusion predominated. Glycerol uptake was decreased by cycloheximide and was more sensitive to sodium azide than to iodoacetate.     

Regulation of phospholipid synthesis in Escherichia coli by guanosine tetraphosphate

Phospholipid synthesis has been reported to be subject to stringent control in Escherichia coli. We present evidence that demonstrates a strict correlation between guanosine tetraphosphate accumulation and inhibition of phospholipid synthesis. In vivo experiments designed to examine the pattern of phospholipid labeling with (32)P-inorganic phosphate and (32)P-sn-glycerol-3-phosphate suggest that regulation must occur at the glycerol-3-phosphate acyltransferase step. Assay of phospholipid synthesis by cell-free extracts and semipurified preparations revealed that guanosine tetraphosphate inhibits at least two enzymes specific for the biosynthetic pathway, sn-glycerol-3-phosphate acyltransferase as well as sn-glycerol-3-phosphate phosphatidyl transferase. These findings provide a biochemical basis for the stringent control of lipid synthesis as well as regulation of steady-state levels of phospholipid in growing cells.     

Enzyme activity profiles in an overwintering population of freeze-tolerant larvae of the gall fly,Eurosta solidaginis

The activity of some enzymes of intermediary metabolism, including enzymes of glycolysis, the hexose monophosphate shunt, and polyol cryoprotectant synthesis, were measured in freeze-tolerant Eurosta solidaginis larvae over a winter season and upon entry into pupation. Flexible metabolic rearrangement was observed concurrently with acclimatization and development. Profiles of enzyme activities related to the metabolism of the cryoprotectant glycerol indicated that fall biosynthesis may occur from two possible pathways: 1. glyceraldehyde-phosphate glyceraldehyde glycerol, using glyceraldehyde phosphatase and NADPH-linked polyol dehydrogenase, or 2. dihydroxyacetonephosphate glycerol-3-phosphate glycerol, using glycerol-3-phosphate dehydrogenase and glycerol-3-phosphatase. Clearance of glycerol in the spring appeared to occur by a novel route through the action of polyol dehydrogenase and glyceraldehyde kinase. Profiles of enzyme activities associated with sorbitol metabolism suggested that this polyol cryoprotectant was synthesized from glucose-6-phosphate through the action of glucose-6-phosphatase and NADPH-linked polyol dehydrogenase. Removal of sorbitol in the spring appeared to occur through the action of sorbitol dehydrogenase and hexokinase. Glycogen phosphorylase activation ensured the required flow of carbon into the synthesis of both glycerol and sorbitol. Little change was seen in the activity of glycolytic or hexose monophosphate shunt enzymes over the winter. Increased activity of the -glycerophosphate shuttle in the spring, indicated by greatly increased glycerol-3-phosphate dehydrogenase activity, may be key to removal and oxidation of reducing equivalents generated from polyol cryoprotectan catabolism.Abbreviations 6PGDH 6-Phosphogluconate dehydrogenase - DHAP dihydroxy acetone phosphate - F6P fructose-6-phosphate - F6Pase fructose-6-phospha-tase - FBPase fructose-bisphosphatase - G3P glycerol-3-phosphate - G3Pase glycerol-3-phosphate phophatase - G3PDH glycerol-3-phosphate dehydrogenase - G6P glucose-6-phosphate - G6Pase glucose-6-phosphatase - G6PDH glucose-6-phosphate dehydrogenase - GAK glyceraldehyde kinase - GAP glyceraldehyde-3-phosphate - GAPase glyceraldehyde-3-phosphatase - GAPDH glyceraldehyde-3-phosphate dehydrogenase - GDH glycerol dehydrogenase - GPase glycogen phosphorylase - HMS hexose monophosphate shunt - LDH lactate dehydrogenase - NADP-IDH NADP⁺-dependent isocitrate dehydrogenase - PDHald polyol dehydrogenase, glyceraldehyde activity - PDHgluc polyol dehydrogenase, glucose activity - PFK phosphofructokinase - PGI phosphoglucoisomerase - PGK phosphoglycerate kinase - PGM phosphoglucomutase - PK pyruvate kinase - PMSF phenylmethylsulfonylfluoride - SoDH sorbitol dehydrogenase - V _max maximal enzyme activity - ww wet weight     

Covalent binding of 3-pyridinealdehyde nicotinamide adenine dinucleotide and substrate to glyceraldehyde 3-phosphate dehydrogenase.

Glyceraldehyde 3-phosphate dehydrogenase (D-glyceraldehyde-3-phoshate:nicotinamide adenine dinucleotide oxidoreductase (phosphorylating), EC 1.2.1.12) forms a complex with 3-pyridinealdehyde-NAD which survives precipitation with 7% perchloric acid. The molar ratio bound 3-pyridinealdehyde-NAD to the enzyme is 2.5 to 2.9. Lactate, malate, and alcohol dehydrogenases do not form acid-precipitable complexes with 3-pyridinealdehyde-NAD. 3-Pyridinealdehyde-deamino-NAD or glyceraldehyde 3-phosphate also forms an acid-stable complex with glyceraldehyde 3-phosphate dehydrogenase; however, NAD, 3-acetylpyridine-NAD, or thionicotinamide-NAD does not produce an acid-stable complex. Incubation of the glyceraldehyde 3-phosphate dehydrogenase with glyceraldehyde 3-phosphate, acetyl phosphate, iodoacetic acid, or iodosobenzoate inhibits the formation of the acid-stable complex with 3-pyridinealdehyde-NAD. Glyceraldehyde 3-phosphate or 3-pyridinealdehyde-NAD also prevents carboxymethylation of the active site cysteine-149 by[14-C]iodoacetic acid. These studies indicate that the aldehyde group of 3-pyridinealdehyde-NAD forms a thiohemiacetal linkage with cysteine-149 which is the substrate binding site for the dehydrogenase reaction. These findings may account for the fact that 3-pyridinealdehyde-NAD strongly inhibits the dehydrogenase and esterase activities of 3-pyridinealdehyde-NAD forms a thiohemiacetal linkage with cysteine-149 which is the substrate binding site for the dehydrogenase reaction. These findings may account for the fact that 3-pyridinealdehyde-NAD strongly inhibits the dehydrogenase and esterase activities of glyceraldehyde 3-phosphate dehydrogenase which require reduced cysteine-149. However, the analogue does not inhibit the acetyl phosphates activity of the enzyme for which the active site sulfhydryl residues must be oxidized.     

The localization of glycerol-3-phosphate dehydrogenase inEscherichia coli

Summary Starved cells ofEscherichia coli are dependent on an exogenous source of energy. It was of interest to ask whether compounds that are commonly used to supply energy must themselves be transported or whether they can be utilized on the outer portion of the cytoplasmic membrane. The utilization of glycerol-3-phosphate an energy source is totally dependent on the membrane-bound glycerol-3-phosphate dehydrogenase. In the present report glycerol-3-phosphate was used as the energy source for uptake of amino acids. A mutant was constructed which is unable to transport this ester and the starved mutant could not drive the uptake of glutamine with glycerol-3-phosphate. It is concluded that the enzyme is located on the internal surface of the membrane in intactE. coli cells. Further evidence was obtained by showing that no glycerol-3-phosphate dehydrogenase activity could be measured in either intact cells or spheroplasts using ferricyanide as electron acceptor, due to its impermeability. The activity could be measured after destruction of the membrane permeability barrier by toluenization. With membrane vesicles prepared according to Kaback's procedure nearly half of the dehydrogenase activity was accessible to ferricyanide as well as to impermeable competitive inhibitors of the enzyme. Partial inversion during preparation of vesicles is the most probable explanation for the results.A protion of this work was presented at the Miami Winter Symposia on the Molecular Basis of Biological Transprot, 1972.     

Pi exchange mediated by the GlpT-dependent sn-glycerol-3-phosphate transport system in Escherichia coli.

The GlpT system for sn-glycerol-3-phosphate transport in Escherichia coli is shown to catalyze a rapid efflux of Pi from the internal phosphate pools in response to externally added Pi or glycerol-3-phosphate. A glpR mutation, which results in constitutive expression of the GlpT system, is responsible for this rapid Pi efflux and the arsenate sensitivity of several laboratory strains, including the popular strain C600. Glucose and other phosphotransferase system sugars inhibit Pi efflux by repressing glpT expression.     

sn-Glycerol-3-phosphate dehydrogenase and its interaction with nitrate reductase in wild-type and hem mutant strains of Staphylococcus aureus

Staphylococcus aureus has membrane-associated sn-glycerol-3-phosphate dehydrogenase activity that is strongly activated by detergents. The enzyme can be measured spectrophotometrically in intact cells in assay systems containing lauryldimethylamine oxide (Ammonyx LO). The dehydrogenase activity was located exclusively in the membrane fraction of cells grown with glycerol under aerobic conditions or under anaerobic conditions with the addition of nitrate; there was no evidence of multiple forms. Development of sn-glycerol-3-phosphate dehydrogenase activity was studied with suspensions of cells grown previously under semianaerobic conditions with glucose and nitrate. The wild-type strain rapidly formed the enzyme when incubated with glycerol under aerobic conditions or under semianaerobic conditions in the presence of nitrate. Under similar conditions, suspensions of hem mutant H-14 required the addition of hemin. Induction of the enzyme was strongly repressed by glucose with both organisms. A procedure was established to obtain cells of mutant H-14 with sn-glycerol-3-phosphate dehydrogenase and nitrate reductase activities, but which could not link the systems unless supplemented with hemin. The coupled activity could also be reconstructed in vitro by the addition of hemin to the depleted membranes.     

Biosynthesis of phosphatidic acid in lipid particles and endoplasmic reticulum of Saccharomyces cerevisiae.

Lipid particles of the yeast Saccharomyces cerevisiae harbor two enzymes that stepwise acylate glycerol-3-phosphate to phosphatidic acid, a key intermediate in lipid biosynthesis. In lipid particles of the s1c1 disruptant YMN5 (M. M. Nagiec et al., J. Biol. Chem. 268:22156-22163, 1993) acylation stops after the first step, resulting in the accumulation of lysophosphatidic acid. Two-dimensional gel electrophoresis confirmed that S1c1p is a component of lipid particles. Lipid particles of a second mutant strain, TTA1 (T. S. Tillman and R. M. Bell, J. Biol. Chem. 261:9144-9149, 1986), which harbors a point mutation in the GAT gene, are essentially devoid of glycerol-3-phosphate acyltransferase activity in vitro. Synthesis of phosphatidic acid is reconstituted by combining lipid particles from YMN5 and TTA1. These results indicate that two distinct enzymes are necessary for phosphatidic acid synthesis in lipid particles: the first step, acylation of glycerol-3-phosphate, is catalyzed by a putative Gat1p; the second step, acylation of lysophosphatidic acid, requires S1c1p. Surprisingly, YMN5 and TTA1 mutants grow like the corresponding wild types because the endoplasmic reticulum of both mutants has the capacity to form a reduced but significant amount of phosphatidic acid. As a consequence, an s1c1 gat1 double mutant is also viable. Lipid particles from this double mutant fail completely to acylate glycerol-3-phosphate, whereas endoplasmic reticulum membranes harbor residual enzyme activities to synthesize phosphatidic acid. Thus, yeast contains at least two independent systems of phosphatidic acid biosynthesis.     

Periplasmic protein related to the sn-glycerol-3-phosphate transport system of Escherichia coli.

Two-dimensional gel electrophoresis of shock fluids of Escherichia coli K-12 revealed the presence of a periplasmic protein related to sn-glycerol-3-phosphate transport (GLPT) that is under the regulation of glpR, the regulatory gene of the glp regulon. Mutants selected for their resistance to phosphonomycin and found to be defective in sn-glycerol-3-phosphate transport either did not produce GLPT or produced it in reduced amounts. Other mutations exhibited no apparent effect of GLPT. Transductions of glpT+ nalA phage P1 into these mutants and selection for growth on sn-glycerol-3-phosphate revealed a 50% cotransduction frequency to nalA. Reversion of mutants taht did not produce GLPT to growth on sn-glycerol-3-phosphate resulted in strains that produce GLPT. This suggests a close relationship of GLPT to the glpT gene and to sn-glycerol-3-phosphate transport. Attempts to demonstrate binding activity of GLPT in crude shock fluid towards sn-glycerol-3-phosphate have failed so far. However, all shock fluids, independent of their GLPT content, exhibited an enzymatic activity that hydrolyzes under the conditions of the binding assay, 30 to 60% of the sn-glycerol-3-phosphate to glycerol and inorganic orthophosphate.     

Characterization of active and latent forms of the membrane-associated sn-glycerol-3-phosphate acyltransferase of Escherichia coli

The intrinsically active, sn-glycerol-3-phosphate acyltransferase present in membranes prepared from both wild type Escherichia coli and from strains which overproduce the enzyme can be kinetically distinguished from a latent enzyme species which is unmasked by solubilization and reconstitution. Both membrane-associated and solubilized/reconstituted enzyme preparations exhibited cooperativity with respect to sn-glycerol-3-phosphate and palmitoyl-coenzyme A substrates; positive cooperativity in membranes toward palmitoyl-coenzyme A (napp = 4) and negative cooperativity toward sn-glycerol-3-phosphate (napp = 0.75) were significantly altered upon solubilization and reconstitution. Since the degree of alteration increased with the amount of sn-glycerol-3-P acyltransferase present in the membranes, a detergent-dissociable homooligomerization of the sn-glycerol-3-phosphate acyltransferase was considered as an underlying mechanism. This possibility was investigated by changing the protein-to-Triton X-100 ratio of homogeneous enzyme prior to reconstitution and then analyzing the subsequent migration of samples on a Sephacryl S-300 sizing column. The elution positions were consistent with monomeric and dimeric polypeptide bound to micelles of Triton X-100. Hill coefficients for monomeric, reconstituted enzyme preparations were comparable to those obtained for the active, membrane-associated sn-glycerol-3-phosphate acyltransferase. The reduced cooperativity of dimeric, reconstituted enzyme preparations correlated closely to the Hill coefficient values obtained for latent, solubilized/reconstituted sn-glycerol-3-phosphate acyltransferase from membranes of Escherichia coli which overproduce the enzyme. The physiological significance of these findings is discussed.     

The initial step of glycerolipid metabolism in Leishmania major promastigotes involves a single glycerol-3-phosphate acyltransferase enzyme important for the synthesis of triacylglycerol but not essential for virulence

The synthesis of the major phospholipids, including those that play an essential role in Leishmania virulence, initiates with the acylation of glycerol-3-phosphate and dihydroxyacetonephosphate at the sn-1 position by glycerol-3-phosphate and dihydroxyacetonephosphate acyltransferases respectively. In this study, we show that Leishmania major promastigotes express a single glycerol-3-phosphate acyltransferase activity important for triacylglycerol synthesis but not essential for virulence. The encoding gene, LmGAT, expressed in yeast results in full complementation of the lethality of a mutant, gat1Deltagat2Delta, lacking glycerol-3-phosphate activity. Biochemical analyses revealed that LmGAT is a low-affinity glycerol-3-phosphate acyltransferase and exhibits higher specific activity with unsaturated long fatty acyl-CoA donors. A L. major null mutant, Deltalmgat/Deltalmgat, was created and a thorough analysis of its lipid composition was performed. Deletion of LmGAT resulted in a complete loss of Leishmania glycerol-3-phosphate acyltransferase activity and a major reduction in triacylglycerol synthesis. Consistent with the specificity of LmGAT for glycerol-3-phosphate but not dihydroxyacetonephosphate, Deltalmgat/Deltalmgat mutant expressed normal levels of the ether-lipid derivatives and virulence factors, lipophosphoglycan and GPI-anchored proteins, gp63, and its virulence was not affected in mice.     

Modification of glyceraldehyde 3-phosphate dehydrogenase activity by adsorption on phospholipid vesicles.

1. The adsorption of [14C]carboxymethylated glyceraldehyde 3-phosphate dehydrogenase to negatively charged liposomes of phsphatidic acid/phosphatidylcholine (3:7, w/w) was investigated. The apparent association constant at I/2 = 60, pH 7.6, was 0.4 X 10(6)M-1. Adsorption decreased as ionic strength and pH were increased. 2. In the presence of negatively charged liposomes, the Km value for glyceraldehyde 3-phosphate of glyceraldehyde 3-phosphate dehydrogenase was increased and Vmax. decreased. In the presence of positively charged liposomes, the Km value for glyceraldehyde 3-phosphate decreased and there was no significant change in Vmax. Addition of Triton X-100 abolished the effect of both positively and negatively charged liposomes on the kinetic properties of the enzyme.     

Intraorganelle localization and substrate specificities of the mitochondrial acyl-CoA: sn-glycerol-3-phosphate O-acyltransferase and acyl-CoA: 1-acyl-sn-glycerol-3-phosphate O-acyltransferase from potato tubers and pea leaves

The mitochondrial sn-glycerol-3-phosphate and 1-acyl-sn-glycerol-3-phosphate O-acyltransferases from potato tubers and pea leaves were investigated with respect to their intraorganelle localization, their positional and substrate specificities, and their fatty acid selectivities. In mitochondria from potato tubers both enzymes were found to be located in the outer membrane. The 1-acyl-sn-glycerol-3-phosphate O-acyltransferase of pea mitochondria showed the same intraorganelle localization whereas the sn-glycerol-3-phosphate O-acyltransferase behaved like a soluble protein of the intermembrane space. The sn-glycerol-3-phosphate O-acyltransferase of both potato and pea mitochondria used sn-glycerol-3-phosphate but not dihydroxyacetone phosphate as acyl acceptor and exclusively catalyzed the formation of 1-acyl-sn-glycerol-3-phosphate which subsequently served as substrate for the second acylation reaction at its C-2 position. Both acyltransferases of potato as well as pea mitochondria showed higher activities with acyl-CoA than with the corresponding acyl-(acyl carrier protein) thioesters. When different acyl-CoA thioesters were offered separately, the sn-glycerol-3-phosphate O-acyltransferase of potato mitochondria displayed no fatty acid specificity whereas the enzyme of pea mitochondria revealed one for saturated acyl groups. On the other hand, the mitochondrial 1-acyl-sn-glycerol-3-phosphate O-acyltransferases from both potato tubers and pea leaves were more active on unsaturated than on saturated acyl-CoA thioesters. Furthermore, these enzymes preferentially used oleoyl- and linoleoyl-CoA when they were offered in a mixture with saturated ones, although the fatty acid selectivity of the pea enzyme was less pronounced than that of the potato enzyme. The sn-glycerol-3-phosphate O-acyltransferase of potato mitochondria displayed a slight preference for saturated acyl groups.     

Relationship between glycerol-3-phosphate dehydrogenase,fatty acid synthase and fatty acid binding proteins in developing human placenta

The activities of the enzymes glycerol-3-phosphate dehydrogenase and fatty acid synthase are inhibited by palmitoyl-coenzyme A and oleate. The two isoforms of fatty acid binding proteins (PI 6.9 and PI 5.4) enhance the activities of glycerol-3-phosphate dehydrogenase and fatty acid synthase in the absence of palmitoyl-coenzyme A or oleate and also protect them against palmitoyl-coenzyme A or oleate inhibition. Levels of fatty acid binding proteins, the activities of the enzymes fatty acid synthase and glycerol-3-phosphate dehydrogenase increase with gestation showing a peak at term. However, the activity of fatty acid synthase showed the same trend up to the 30th week of gestation and then declined slightly at term. With the advancement of pregnancy when more lipids are required for the developing placenta, fatty acid binding proteins supply more fatty acids and glycerol-3-phosphate for the synthesis of lipids. Thus a correlation exists between glycerol-3-phosphate dehydrogenase, fatty acid synthase and fatty acid binding proteins in developing human placenta.     

sn-glycerol-3-phosphate oxidase and alcohol tolerance inDrosophila melanogaster larvae

The role of sn-glycerol-3-phosphate oxidase (GPO; EC 1.1.99.5) in the variation of ethanol tolerance in Drosophila melanogaster was assessed in isofemale lines derived from individuals collected at the Chateau Tahbilk Winery and Wandin North Orchard of Victoria, Australia. When fed an undefined medium (semolina-treacle) with 6% ethanol (v/v), larvae of lines with high GPO activities survived better than did larvae of lines with low GPO activities. Although GPO was induced to higher activity levels by dietary ethanol in larvae of all the test lines, GPO activity was greater in lines representing the area outside the wine cellar. This implied that the cellar environment selected against individuals with high levels of GPO. These data do not explain the established difference in tolerance between cellar and outside populations. The GPO activities of lines were not dependent upon the activities of the lipogenic enzyme, glycerol-3-phosphate dehydrogenase; the major ethanol-degrading enzyme, alcohol dehydrogenase; or the citric acid cycle enzyme, fumarase. Thus, GPO activity is an important component of the metabolic mechanism of ethanol tolerance in larvae, but the mode of action of GPO has not been defined.