| 1. |
- Alshahawy, Rasha, et al.
(författare)
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Changes in plasma fatty acids and related biomarkers during transition to an exclusively plant- and fish-based diet in healthy adults
- 2021
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Ingår i: Nutrition (Burbank, Los Angeles County, Calif.). - : Elsevier. - 0899-9007 .- 1873-1244. ; 90
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Tidskriftsartikel (refereegranskat)abstract
- Objective: The aim of this study was to examine the time scale of plasma fatty acid changes during transition to an exclusively plant- and fish-based diet in healthy individuals and determine whether there are associated alterations in arachidonic acid (ARA)-derived inflammatory mediators, estimated stearoyl coenzyme A desaturase (SCD) activity, and blood pressure. Methods: In pursuit of a religious fast, 36 adults abstained from eating poultry, meat, dairy products, and eggs, while increasing fish intake for 6 wk. Participants were assessed 1 wk before (W0) and 1 (WI) and 6 (W6) weeks after the diet change. Results: By W6, fasting plasma long-chain omega-3 polyunsaturated fatty acids (omega-3 LC-PUFAs); docosahexaenoic (DHA) and eicosapentaenoic (EPA) had increased (+67% and +73%, respectively; P <= 0.001), with early rise of DHA (+22%), but not EPA at W1.The omega-3 index (sum of DHA and EPA as a percent of total fatty acids) increased from 2.1% to 3.4%. ARA decreased progressively (W1, -9%; W6, -16%; P < 0.001). ARA precursors gamma-linolenic and dihomo-gamma-linolenic acids also decreased, without changes in the ARA-derived mediators prostaglandin-E2 and leukotriene-B4. Myristic acid decreased at W1 (-37%) and W6 (-40%). There was no consistent change in SCD indices. At W6, systolic and diastolic blood pressure had declined by 8 and 5 mm Hg, respectively (P <= 0.013). Conclusions: Shifting to a plant- and fish-based diet produces rapid and sustained increases in omega-3 LC-PUFAs and decreases the omega-6 PUFA ARA and its precursors, consistent with a cardio-protective profile. The rapid response suggests that these biomarkers may be useful for assessment of diet interventions.
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| 2. |
- Chen, Yang, et al.
(författare)
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Purification and site-directed mutagenesis of linoleate 9S-dioxygenase-allene oxide synthase of Fusarium oxysporum confirms the oxygenation mechanism
- 2017
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Ingår i: Archives of Biochemistry and Biophysics. - : ELSEVIER SCIENCE INC. - 0003-9861 .- 1096-0384. ; 625-626, s. 24-29
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Tidskriftsartikel (refereegranskat)abstract
- Plants and fungi form jasmonic acid from a-linolenic acid. The first two steps of biosynthesis in plants occur by sequential transformation by 13S-Iipoxygenase and allene oxide synthase (AOS). The biosynthesis in fungi may follow this classical scheme, but the only fungal AOS discovered so far are cytochromes P450 (CYP) fused to 8- and 9-dioxygenases (DOX). In the present report, we purified recombinant 9S-DOX-AOS of Fusarium oxysporum from cell lysate by cobalt affinity chromatography to near homogeneity and studied key residues by site-directed mutagenesis. Sequence homology with 8R-DOX-linoleate diol synthases (8R-DOX-LDS) suggested that Tyr414 catalyzes hydrogen abstraction and that Cys1051 forms the heme thiolate ligand. Site-directed mutagenesis (Tyr414Phe; Cys1051Ser) led to loss of 9S-DOX and 9S-AOS activities, respectively, but other important residues in the CYP parts of 5,8 and 7,8-LDS or 9R-AOS were not conserved. The UV-visible spectrum of 9S-DOX-AOS showed a Soret band at 409 nm, which shifted to 413 nm in the Cys1051Ser mutant. The 9S-AOS of the Tyr414Phe mutant transformed 9S-hydroperoxides of alpha-linolenic and linoleic acids to allene oxides/alpha-ketols, but it did not transform 13-hydroperoxides. We conclude that 9S- and 8R-DOX catalyze hydrogen abstraction at C-11 and C-8, respectively, by homologous Tyr residues.
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| 3. |
- Elkafrawy, Hagar, et al.
(författare)
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Extracellular cystine influences human preadipocyte differentiation and correlates with fat mass in healthy adults
- 2021
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Ingår i: Amino Acids. - : Springer Nature. - 0939-4451 .- 1438-2199. ; 53, s. 1623-1634
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Tidskriftsartikel (refereegranskat)abstract
- Plasma cysteine is associated with human obesity, but it is unknown whether this is mediated by reduced, disulfide (cystine and mixed-disulfides) or protein-bound (bCys) fractions. We investigated which cysteine fractions are associated with adiposity in vivo and if a relevant fraction influences human adipogenesis in vitro. In the current study, plasma cysteine fractions were correlated with body fat mass in 35 adults. Strong positive correlations with fat mass were observed for cystine and mixed disulfides (r >= 0.61, P < 0.001), but not the quantitatively major form, bCys. Primary human preadipocytes were differentiated in media containing cystine concentrations varying from 10-50 mu M, a range similar to that in plasma. Increasing extracellular cystine (10-50 mu M) enhanced mRNA expression of PPARG2 (to sixfold), PPARG1, PLIN1, SCD1 and CDO1 (P = 0.042- < 0.001). Adipocyte lipid accumulation and lipid-droplet size showed dose-dependent increases from lowest to highest cystine concentrations (P < 0.001), and the malonedialdehyde/total antioxidant capacity increased, suggesting increased oxidative stress. In conclusion, increased cystine concentrations, within the physiological range, are positively associated with both fat mass in healthy adults and human adipogenic differentiation in vitro. The potential role of cystine as a modifiable factor regulating human adipocyte turnover and metabolism deserves further study.
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| 4. |
- Elshorbagy, Amany, et al.
(författare)
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Amino acid changes during transition to a vegan diet supplemented with fish in healthy humans.
- 2017
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Ingår i: European Journal of Nutrition. - : Springer Science and Business Media LLC. - 1436-6207 .- 1436-6215. ; 56:5, s. 1953-1962
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Tidskriftsartikel (refereegranskat)abstract
- PURPOSE: To explore whether changes in dietary protein sources can lower plasma branched-chain amino acids (BCAAs), aromatic amino acids and sulfur amino acids (SAAs) that are often elevated in the obese, insulin-resistant state and in type 2 diabetes.METHODS: Thirty-six subjects (mean age 31 ± 2 years) underwent a voluntary abstinence from meat, poultry, eggs, and dairy products for 6 weeks, while enriching the diet with fish, in fulfillment of a religious fast. Subjects were assessed 1 week before the fast (V1), 1 week after initiation of the fast (V2) and in the last week of the fast (V3). Thirty-four subjects completed all three visits.RESULTS: Fasting plasma BCAAs decreased at V2 and remained low at V3 (P < 0.001 for all). Valine showed the greatest decline, by 20 and 19 % at V2 and V3, respectively. Phenylalanine and tryptophan, but not tyrosine, also decreased at V2 and V3. The two proteinogenic SAAs, methionine and cysteine, remained stable, but the cysteine product, taurine, decreased from 92 ± 7 μmol/L to 66 ± 6 (V2; P = 0.003) and 65 ± 6 μmol/L (V3; P = 0.003). A progressive decline in plasma glutamic acid, coupled with an increase in glutamine, was observed. Plasma total and LDL cholesterol decreased at V2 and V3 (P < 0.001 for all).CONCLUSION: Changing dietary protein sources to plant- and fish-based sources in an ad libitum setting lowers the plasma BCAAs that have been linked to diabetes risk. These findings point to habitual diet as a potentially modifiable determinant of fasting plasma BCAA concentrations.
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| 5. |
- Elshorbagy, Amany K, et al.
(författare)
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Exploring the Lean Phenotype of Glutathione-Depleted Mice : Thiol, Amino Acid and Fatty Acid Profiles.
- 2016
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 11:10
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Although reduced glutathione (rGSH) is decreased in obese mice and humans, block of GSH synthesis by buthionine sulfoximine (BSO) results in a lean, insulin-sensitive phenotype. Data is lacking about the effect of BSO on GSH precursors, cysteine and glutamate. Plasma total cysteine (tCys) is positively associated with stearoyl-coenzyme A desaturase (SCD) activity and adiposity in humans and animal models.OBJECTIVE: To explore the phenotype, amino acid and fatty acid profiles in BSO-treated mice.DESIGN: Male C3H/HeH mice aged 11 weeks were fed a high-fat diet with or without BSO in drinking water (30 mmol/L) for 8 weeks. Amino acid and fatty acid changes were assessed, as well as food consumption, energy expenditure, locomotor activity, body composition and liver vacuolation (steatosis).RESULTS: Despite higher food intake, BSO decreased particularly fat mass but also lean mass (both P<0.001), and prevented fatty liver vacuolation. Physical activity increased during the dark phase. BSO decreased plasma free fatty acids and enhanced insulin sensitivity. BSO did not alter liver rGSH, but decreased plasma total GSH (tGSH) and rGSH (by ~70%), and liver tGSH (by 82%). Glutamate accumulated in plasma and liver. Urine excretion of cysteine and its precursors was increased by BSO. tCys, rCys and cystine decreased in plasma (by 23-45%, P<0.001 for all), but were maintained in liver, at the expense of decreased taurine. Free and total plasma concentrations of the SCD products, oleic and palmitoleic acids were decreased (by 27-38%, P <0.001 for all).CONCLUSION: Counterintuitively, block of GSH synthesis decreases circulating tCys, raising the question of whether the BSO-induced obesity-resistance is linked to cysteine depletion. Cysteine-supplementation of BSO-treated mice is warranted to dissect the effects of cysteine and GSH depletion on energy metabolism.
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| 6. |
- Elshorbagy, Amany K., et al.
(författare)
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Food Overconsumption in Healthy Adults Triggers Early and Sustained Increases in Serum Branched-Chain Amino Acids and Changes in Cysteine Linked to Fat Gain
- 2018
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Ingår i: Journal of Nutrition. - : OXFORD UNIV PRESS. - 0022-3166 .- 1541-6100. ; 148:7, s. 1073-1080
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Tidskriftsartikel (refereegranskat)abstract
- Background: Plasma concentrations of branched-chain amino acids (BCAAs) and the sulfur-containing amino acid cysteine are associated with obesity and insulin resistance. BCAAs predict future diabetes. Objective: We investigated amino acid changes during food overconsumption. Methods: Forty healthy men and women with a body mass index (mean +/- SEM) of 25.6 +/- 0.6 were overfed by 1250 kcal/d for 28 d, increasing consumption of all macronutrients. Insulin sensitivity and body composition were assessed at baseline (day 0) and day 28. Fasting serum amino acids were measured at days 0, 3, and 28. Linear mixed-effects models evaluated the effect of time in the total group and separately in those with low and high body fat gain (below compared with at or above median fat gain, 1.95 kg). At days 0 and 28, insulin-induced suppression of serum amino acids during a hyperinsulinemic-euglycemic clamp test and, in a subset (n = 20), adipose tissue mRNA expression of selected amino acid metabolizing enzymes were assessed. Results: Weight increased by 2.8 kg. High fat gainers gained 2.6 kg fat mass compared with 1.1 kg in low fat gainers. Valine and isoleucine increased at day 3 (+17% and +22%, respectively; P <= 0.002) and remained elevated at day 28, despite a decline in valine (P = 0.019) from day 3 values. Methionine, cystathionine, and taurine were unaffected. Serum total cysteine (tCys) transiently increased at day 3 (+11%; P = 0.022) only in high fat gainers (P-interaction = 0.043), in whom the cysteine catabolic enzyme cysteine dioxygenase (CDO1) was induced (+26%; P = 0.025) in adipose tissue (P-interaction = 0.045). Overconsumption did not alter adipose tissue mRNA expression of the BCAA-metabolizing enzymes branched-chain keto acid dehydrogenase E1 alpha polypeptide (BCKDHA) or branched-chain amino transferase 1 (BCAT1). In the total population at day 0, insulin infusion decreased all serum amino acids (-11% to -47%; P < 0.01), except for homocysteine and tCys, which were unchanged, and glutathione, which was increased by 54%. At day 28, insulin increased tCys (+8%), and the insulin-induced suppression of taurine and phenylalanine observed at day 0, but not that of BCAAs, was significantly impaired. Conclusions: These findings highlight the role of nutrient oversupply in increasing fasting BCAA concentrations in healthy adults. The link between cysteine availability, CDO1 expression, and fat gain deserves investigation.
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| 7. |
- Elshorbagy, A K, et al.
(författare)
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Serum S-adenosylmethionine, but not methionine, increases in response to overfeeding in humans.
- 2016
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Ingår i: Nutrition & Diabetes. - : Springer Science and Business Media LLC. - 2044-4052. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Plasma concentration of the methyl donor S-adenosylmethionine (SAM) is linearly associated with body mass index (BMI) and fat mass. As SAM is a high-energy compound and a sensor of cellular nutrient status, we hypothesized that SAM would increase with overfeeding.METHODS: Forty normal to overweight men and women were overfed by 1250 kcal per day for 28 days.RESULTS: Serum SAM increased from 106 to 130 nmol/l (P=0.006). In stratified analysis, only those with weight gain above the median (high-weight gainers; average weight gain 3.9±0.3 kg) had increased SAM (+42%, P=0.001), whereas low-weight gainers (weight gain 1.5±0.2 kg) did not (Pinteraction=0.018). Overfeeding did not alter serum concentrations of the SAM precursor, methionine or the products, S-adenosyl-homocysteine and homocysteine. The SAM/SAH (S-adenosylhomocysteine) ratio was unchanged in the total population, but increased in high-weight gainers (+52%, P=0.006, Pinteraction =0.005). Change in SAM correlated positively with change in weight (r=0.33, P=0.041) and fat mass (r=0.44, P=0.009), but not with change in protein intake or plasma methionine, glucose, insulin or low-density lipoprotein (LDL)-cholesterol.CONCLUSION: Overfeeding raised serum SAM in proportion to the fat mass gained. The increase in SAM may help stabilize methionine levels, and denotes a responsiveness of SAM to nutrient state in humans. The role of SAM in human energy metabolism deserves further attention.
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| 8. |
- Garscha, Ulrike, et al.
(författare)
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Identification of dioxygenases required for Aspergillus development : Studies of products, stereochemistry, and the reaction mechanism
- 2007
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 282:48, s. 34707-34718
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Tidskriftsartikel (refereegranskat)abstract
- Aspergillus sp. contain ppoA, ppoB, and ppoC genes, which code for fatty acid oxygenases with homology to fungal linoleate 7,8-diol synthases (7,8-LDS) and cyclooxygenases. Our objective was to identify these enzymes, as ppo gene replacements show critical developmental aberrancies in sporulation and pathogenicity in the human pathogen Aspergillus fumigatus and the genetic model Aspergillus nidulans. The PpoAs of A. fumigatus and A. nidulans were identified as (8R)-dioxygenases with hydroperoxide isomerase activity, designated 5,8-LDS. 5,8-LDS transformed 18:2n-6 to (8R)-hydroperoxyoctadecadienoic acid ((8R)-HPODE) and (5S,8R)-dihydroxy-9Z,12Z-octadecadienoic acid ((5S,8R)-DiHODE). We also detected 8,11-LDS in A. fumigatus and (10R)-dioxygenases in both Aspergilli. The diol synthases oxidized [(8R)-2H]18:2n-6 to (8R)-HPODE with retention of the deuterium label, suggesting antarafacial hydrogen abstraction and insertion of molecular oxygen. Experiments with stereospecifically deuterated 18:2n-6 showed that (8R)-HPODE was isomerized by 5,8- and 8,11-LDS to (5S,8R)-DiHODE and to (8R,11S)-dihydroxy-9Z,12Z-octadecadienoic acid, respectively, by suprafacial hydrogen abstraction and oxygen insertion at C-5 and C-11. PpoCs were identified as (10R)-dioxygenases, which catalyzed abstraction of the pro-S hydrogen at C-8 of 18:2n-6, double bond migration, and antafacial insertion of molecular oxygen with formation of (10R)-hydroxy-8E,12Z- hydroperoxyoctadecadienoic acid ((10R)-HPODE). Deletion of ppoA led to prominent reduction of (8R)-H(P)ODE and complete loss of (5S,8R)-DiHODE biosynthesis, whereas biosynthesis of (10R)-HPODE was unaffected. Deletion of ppoC caused biosynthesis of traces of racemic 10-HODE but did not affect the biosynthesis of other oxylipins. We conclude that ppoA of Aspergillus sp. may code for 5,8-LDS with catalytic similarities to 7,8-LDS and ppoC for linoleate (10R)-dioxygenases. Identification of these oxygenases and their products will provide tools for analyzing the biological impact of oxylipin biosynthesis in Aspergilli.
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| 9. |
- Haj-Yasein, Nadia Nabil, et al.
(författare)
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Cysteine deprivation prevents induction of peroxisome proliferator-activated receptor gamma-2 and adipose differentiation of 3T3-L1 cells.
- 2017
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Ingår i: Biochimica et Biophysica Acta. - : Elsevier BV. - 0006-3002 .- 1878-2434. ; 1862:6, s. 623-635
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Tidskriftsartikel (refereegranskat)abstract
- Plasma cysteine is strongly associated with body fat mass in human cohorts and diets low in cysteine prevents fat accumulation in mice. It is unclear if plasma cysteine affects fat development or if fat accumulation raises plasma cysteine. To determine if cysteine affects adipogenesis, we differentiated 3T3-L1 preadipocytes in medium with reduced cysteine. Cells incubated in media with 10-20μM cysteine exhibited reduced capacity to differentiate into triacylglycerol-storing mature adipocytes compared with cells incubated with 50μM cysteine. Low cysteine severely reduced expression of peroxisome proliferator-activated receptor gamma2 (Pparγ2) and its target genes perlipin1 (Plin1) and fatty acid binding protein-4 (Fabp4). Expression of stearoyl-CoA desaturase-1 (Scd1), known to be repressed with cysteine depletion, was also reduced with low cysteine. Medium depletion of the essential amino acids leucine, valine, and isoleucine had only a modest effect on adipocyte specific gene expression and differentiation. Stimulation with the PPARγ agonist BRL-49653 or addition of a hydrogen sulfide donor enhanced differentiation of 3T3-L1 cells cultured in low cysteine. This demonstrates that the ability to induce PPARγ expression is preserved when cells are cultured in low cysteine. It therefore appears that cysteine depletion inhibits adipogenesis by specifically affecting molecular pathways required for induction of PPARγ expression, rather than through a general reduction of global protein synthesis. In conclusion, we show that low extracellular cysteine reduces adipocyte differentiation by interfering with PPARγ2 and PPARγ target gene expression. Our results provide further evidence for the hypothesis that plasma cysteine is a casual determinant for body fat mass.
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| 10. |
- Hoffmann, Inga, et al.
(författare)
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Epoxy alcohol synthase of the rice blast fungus represents a novel subfamily of dioxygenase-cytochrome P450 fusion enzymes
- 2014
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Ingår i: Journal of Lipid Research. - 0022-2275 .- 1539-7262. ; 55:10, s. 2113-2123
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Tidskriftsartikel (refereegranskat)abstract
- The genome of the rice blast fungus Magnaporthe oryzae codes for two proteins with N-terminal dioxygenase (DOX) and C-terminal cytochrome P450 (CYP) domains, respectively. One of them, MGG_ 13239, was confirmed as 7,8-linoleate diol synthase by prokaryotic expression. The other recombinant protein (MGG_ 10859) possessed prominent 10R-DOX and epoxy alcohol synthase (EAS) activities. This enzyme, 10R-DOX-EAS, transformed 18:2n-6 sequentially to 10(R)-hydroperoxy-8(E), 12(Z)-octadecadienoic acid (10R-HPODE) and to 12S(13R)-epoxy-10(R)-hydroxy-8(E)octadecenoic acid as the end product. Oxygenation at C-10 occurred by retention of the pro-R hydrogen of C-8 of 18:2n-6, suggesting antarafacial hydrogen abstraction and oxygenation. Experiments with O-18(2) and O-16(2) gas confirmed that the epoxy alcohol was formed from 10R-HPODE, likely by heterolytic cleavage of the dioxygen bond with formation of P450 compound I, and subsequent intramolecular epoxidation of the 12(Z) double bond. Site-directed mutagenesis demonstrated that the cysteinyl heme ligand of the P450 domain was required for the EAS activity. Replacement of Asn(965) with Val in the conserved AsnGlnXaaGln sequence revealed that Asn965 supported formation of the epoxy alcohol. 10R-DOX-EAS is the first member of a novel subfamily of DOX-CYP fusion proteins of devastating plant pathogens.
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