Flexible B12 ecophysiology of Phaeocystis antarctica due to a fusion B12-independent methionine synthase with widespread homologues.

Coastal Antarctic marine ecosystems are significant in carbon cycling because of their intense seasonal phytoplankton blooms. Southern Ocean algae are primarily limited by light and iron (Fe) and can be co-limited by cobalamin (vitamin B12). Micronutrient limitation controls productivity and shapes the composition of blooms which are typically dominated by either diatoms or the haptophyte Phaeocystis antarctica. However, the vitamin requirements and ecophysiology of the keystone species P. antarctica remain poorly characterized. Using cultures, physiological analysis, and comparative omics, we examined the response of P. antarctica to a matrix of Fe-B12 conditions. We show that P. antarctica is not auxotrophic for B12, as previously suggested, and identify mechanisms underlying its B12 response in cultures of predominantly solitary and colonial cells. A combination of proteomics and proteogenomics reveals a B12-independent methionine synthase fusion protein (MetE-fusion) that is expressed under vitamin limitation and interreplaced with the B12-dependent isoform under replete conditions. Database searches return homologues of the MetE-fusion protein in multiple Phaeocystis species and in a wide range of marine microbes, including other photosynthetic eukaryotes with polymorphic life cycles as well as bacterioplankton. Furthermore, we find MetE-fusion homologues expressed in metaproteomic and metatranscriptomic field samples in polar and more geographically widespread regions. As climate change impacts micronutrient availability in the coastal Southern Ocean, our finding that P. antarctica has a flexible B12 metabolism has implications for its relative fitness compared to B12-auxotrophic diatoms and for the detection of B12-stress in a more diverse set of marine microbes.

Growth requirement for methionine in human melanoma-derived cell lines with different levels of MMACHC expression and methylation.

Methionine dependence, the inability to grow in culture when methionine in the medium is replaced by its metabolic precursor homocysteine, occurs in many tumor cell lines. In most affected lines, the cause of methionine dependence is not known. An exception is the melanoma-derived cell line MeWo-LC1, in which hypermethylation of the MMACHC gene is associated with decreased MMACHC expression. Decreased expression results in decreased provision of the methylcobalamin cofactor required for activity of methionine synthase and thus decreased conversion of homocysteine to methionine. Analysis of data in the Cancer Cell Line Encyclopedia Archive demonstrated that MMACHC hypermethylation and decreased MMACHC expression occurred more frequently in melanoma cell lines when compared to other tumor cell lines. We further investigated methionine dependence and aspects of MMACHC function in a panel of six melanoma lines, including both melanoma lines with known methionine dependence status (MeWo, which is methionine independent, and A375, which is methionine dependent). We found that the previously unclassified melanoma lines HMCB, Colo829 and SH-4 were methionine dependent, while SK-Mel-28 was methionine independent. However, despite varying levels of MMACHC methylation and expression, none of the tested lines had decreased methylcobalamin and adenosylcobalamin synthesis as seen in MeWo-LC1, and the functions of both cobalamin-dependent enzymes methionine synthase and methylmalonyl-CoA mutase were intact. Thus, while melanoma lines were characterized by relatively high levels of MMACHC methylation and low expression, the defect in metabolism observed in MeWo-LC1 was unique, and decreased MMACHC expression was not a cause of methionine dependence in the other melanoma lines.

Structure of full-length cobalamin-dependent methionine synthase and cofactor loading captured in crystallo.

Cobalamin-dependent methionine synthase (MS) is a key enzyme in methionine and folate one-carbon metabolism. MS is a large multi-domain protein capable of binding and activating three substrates: homocysteine, folate, and S-adenosylmethionine for methylation. Achieving three chemically distinct methylations necessitates significant domain rearrangements to facilitate substrate access to the cobalamin cofactor at the right time. The distinct conformations required for each reaction have eluded structural characterization as its inherently dynamic nature renders structural studies difficult. Here, we use a thermophilic MS homolog (tMS) as a functional MS model. Its exceptional stability enabled characterization of MS in the absence of cobalamin, marking the only studies of a cobalamin-binding protein in its apoenzyme state. More importantly, we report the high-resolution full-length MS structure, ending a multi-decade quest. We also capture cobalamin loading in crystallo, providing structural insights into holoenzyme formation. Our work paves the way for unraveling how MS orchestrates large-scale domain rearrangements crucial for achieving challenging chemistries.

The role of methionine synthases in fungal metabolism and virulence.

Methionine synthases (MetH) catalyse the methylation of homocysteine (Hcy) with 5-methyl-tetrahydrofolate (5, methyl-THF) acting as methyl donor, to form methionine (Met) and tetrahydrofolate (THF). This function is performed by two unrelated classes of enzymes that differ significantly in both their structures and mechanisms of action. The genomes of plants and many fungi exclusively encode cobalamin-independent enzymes (EC.2.1.1.14), while some fungi also possess proteins from the cobalamin-dependent (EC.2.1.1.13) family utilised by humans. Methionine synthase's function connects the methionine and folate cycles, making it a crucial node in primary metabolism, with impacts on important cellular processes such as anabolism, growth and synthesis of proteins, polyamines, nucleotides and lipids. As a result, MetHs are vital for the viability or virulence of numerous prominent human and plant pathogenic fungi and have been proposed as promising broad-spectrum antifungal drug targets. This review provides a summary of the relevance of methionine synthases to fungal metabolism, their potential as antifungal drug targets and insights into the structures of both classes of MetH.

Conformational switching and flexibility in cobalamin-dependent methionine synthase studied by small-angle X-ray scattering and cryoelectron microscopy.

Cobalamin-dependent methionine synthase (MetH) catalyzes the synthesis of methionine from homocysteine and 5-methyltetrahydrofolate (CH3-H4folate) using the unique chemistry of its cofactor. In doing so, MetH links the cycling of S-adenosylmethionine with the folate cycle in one-carbon metabolism. Extensive biochemical and structural studies on Escherichia coli MetH have shown that this flexible, multidomain enzyme adopts two major conformations to prevent a futile cycle of methionine production and consumption. However, as MetH is highly dynamic as well as both a photosensitive and oxygen-sensitive metalloenzyme, it poses special challenges for structural studies, and existing structures have necessarily come from a "divide and conquer" approach. In this study, we investigate E. coli MetH and a thermophilic homolog from Thermus filiformis using small-angle X-ray scattering (SAXS), single-particle cryoelectron microscopy (cryo-EM), and extensive analysis of the AlphaFold2 database to present a structural description of the full-length MetH in its entirety. Using SAXS, we describe a common resting-state conformation shared by both active and inactive oxidation states of MetH and the roles of CH3-H4folate and flavodoxin in initiating turnover and reactivation. By combining SAXS with a 3.6-Å cryo-EM structure of the T. filiformis MetH, we show that the resting-state conformation consists of a stable arrangement of the catalytic domains that is linked to a highly mobile reactivation domain. Finally, by combining AlphaFold2-guided sequence analysis and our experimental findings, we propose a general model for functional switching in MetH.

Cognitive Impairment Is Associated with AMPAR Glutamatergic Dysfunction in a Mouse Model of Neuronal Methionine Synthase Deficiency.

Impairment of one-carbon metabolism during pregnancy, either due to nutritional deficiencies in B9 or B12 vitamins or caused by specific genetic defects, is often associated with neurological defects, including cognitive dysfunction that persists even after vitamin supplementation. Animal nutritional models do not allow for conclusions regarding the specific brain mechanisms that may be modulated by systemic compensations. Using the Cre-lox system associated to the neuronal promoter Thy1.2, a knock-out model for the methionine synthase specifically in the brain was generated. Our results on the neurobehavioral development of offspring show that the absence of methionine synthase did not lead to growth retardation, despite an effective reduction of both its expression and the methylation status in brain tissues. Behaviors were differently affected according to their functional outcome. Only temporary retardations were recorded in the acquisition of vegetative functions during the suckling period, compared to a dramatic reduction in cognitive performance after weaning. Investigation of the glutamatergic synapses in cognitive areas showed a reduction of AMPA receptors phosphorylation and clustering, indicating an epigenomic effect of the neuronal deficiency of methionine synthase on the reduction of glutamatergic synapses excitability. Altogether, our data indicate that cognitive impairment associated with methionine synthase deficiency may not only result from neurodevelopmental abnormalities, but may also be the consequence of alterations in functional plasticity of the brain.

Development of vitamin B12 dependency in Saccharomyces cerevisiae.

For decades, the industrial vitamin B12 (cobalamin) production has been based on bacterial producer strains. Due to limited methods for strain optimization and difficult strain handling, the desire for new vitamin B12-producing hosts has risen. As a vitamin B12-independent organism with a big toolbox for genomic engineering and easy-to-handle cultivation conditions, Saccharomyces cerevisiae has high potential for heterologous vitamin B12 production. However, the B12 synthesis pathway is long and complex. To be able to easily engineer and evolve B12-producing recombinant yeast cells, we have developed an S. cerevisiae strain whose growth is dependent on vitamin B12. For this, the B12-independent methionine synthase Met6 of yeast was replaced by a B12-dependent methionine synthase MetH from Escherichia coli. Adaptive laboratory evolution, RT-qPCR, and overexpression experiments show that additional high-level expression of a bacterial flavodoxin/ferredoxin-NADP+ reductase (Fpr-FldA) system is essential for in vivo reactivation of MetH activity and growth. Growth of MetH-containing yeast cells on methionine-free media is only possible with the addition of adenosylcobalamin or methylcobalamin. A heterologous vitamin B12 transport system turned out to be not necessary for the uptake of cobalamins. This strain should be a powerful chassis to engineer B12-producing yeast cells.

Legionellapneumophila induces methylomic changes in ten-eleven translocation to ensure bacterial reproduction in human lung epithelial cells.

Introduction. Legionella pneumophila is a Gram-negative flagellated bacteria that can infect human lungs and cause a severe form of pneumonia named Legionnaires' disease.Hypothesis. We hypothesize that L. pneumophila infection induces methylomic changes in methylcytosine dioxygenases, ten-eleven translocation (TET) genes, and controls DNA methylation following infection.Aim. In the current research, we sought to further investigate DNA methylation changes in human lung epithelial cells upon L. pneumophila infection and determine how methylation inhibitor agents disturb L. pneumophila reproduction.Methodology. A549 cell line was used in L. pneumophila infection and inhibitors' treatment, including 5-azacytidine (5-AZA) and (-)-epigallocatechin-3-O-gallate (EGCG).Results. Interestingly, DNA methylation analysis of infected A549 using sodium bisulfite PCR and the methylation-sensitive HpaII enzyme showed potential methylation activity within the promoter regions of ten-eleven translocation (TET) genes located on CpG/397-8 and CpG/385-6 of TET1 and TET3, respectively. Such methylation changes in TET effectors decreased their expression profile following infection, indicated by quantitative real-time PCR (RT-qPCR), immunoblotting and flow cytometry. Furthermore, pre-treatment of A549 cells with 5-AZA or EGCG significantly decreased the bacterial reproduction characterized by the expression of L. pneumophila 16S ribosomal RNA and the c.f.u. ml-1 of bacterial particles. Moreover, both methylation inhibitors showed potent inhibition of methionine synthase (MS) expression, which was further confirmed by the docking analysis of inhibitor ligands and crystal structure of MS protein.Conclusion. These data provide evidence for the methylomic changes in the promoter region of TET1 and TET3 by L. pneumophila infection in the A549 cell line and suggest the anti-bacterial properties of 5-AZA and EGCG, as methylation inhibitors, are due to targeting the epigenetic effector methionine synthase.

SAMS-1 coordinates HLH-30/TFEB and PHA-4/FOXA activities through histone methylation to mediate dietary restriction-induced autophagy and longevity.

Dietary restriction (DR) is known to promote autophagy to exert its longevity effect. While SAMS-1 (S-adenosyl methionine synthetase-1) has been shown to be a key mediator of the DR response, little is known about the roles of S-adenosyl methionine (SAM) and SAM-dependent methyltransferase in autophagy and DR-induced longevity. In this study, we show that DR and SAMS-1 repress the activity of SET-2, a histone H3K4 methyltransferase, by limiting the availability of SAM. Consequently, the reduced H3K4me3 levels promote the expression and activity of two transcription factors, HLH-30/TFEB and PHA-4/FOXA, which both regulate the transcription of autophagy-related genes. We then find that HLH-30/TFEB and PHA-4/FOXA act collaboratively on their common target genes to mediate the transcriptional response of autophagy-related genes and consequently the lifespan of the animals. Our study thus shows that the SAMS-1-SET-2 axis serves as a nutrient-sensing module to epigenetically coordinate the activation of HLH-30/TFEB and PHA-4/FOXA transcription factors to control macroautophagy/autophagy and longevity in response to DR.Abbreviations: ChIP: chromatin immunoprecipitation; ChIP-seq: chromatin immuno precipitation-sequencing; COMPASS: complex of proteins associated with Set1; DR: dietary restriction; GO: gene ontology; SAM: S-adenosyl methionine; SAMS-1: S-adenosyl methionine synthetase-1; TSS: transcription start site; WT: wild-type.

High Folate, Perturbed One-Carbon Metabolism and Gestational Diabetes Mellitus.

Folate is a dietary micronutrient essential to one-carbon metabolism. The World Health Organisation recommends folic acid (FA) supplementation pre-conception and in early pregnancy to reduce the risk of fetal neural tube defects (NTDs). Subsequently, many countries (~92) have mandatory FA fortification policies, as well as recommendations for periconceptional FA supplementation. Mandatory fortification initiatives have been largely successful in reducing the incidence of NTDs. However, humans have limited capacity to incorporate FA into the one-carbon metabolic pathway, resulting in the increasingly ubiquitous presence of circulating unmetabolised folic acid (uFA). Excess FA intake has emerged as a risk factor in gestational diabetes mellitus (GDM). Several other one-carbon metabolism components (vitamin B12, homocysteine and choline-derived betaine) are also closely entwined with GDM risk, suggesting a role for one-carbon metabolism in GDM pathogenesis. There is growing evidence from in vitro and animal studies suggesting a role for excess FA in dysregulation of one-carbon metabolism. Specifically, high levels of FA reduce methylenetetrahydrofolate reductase (MTHFR) activity, dysregulate the balance of thymidylate synthase (TS) and methionine synthase (MTR) activity, and elevate homocysteine. High homocysteine is associated with increased oxidative stress and trophoblast apoptosis and reduced human chorionic gonadotrophin (hCG) secretion and pancreatic ß-cell function. While the relationship between high FA, perturbed one-carbon metabolism and GDM pathogenesis is not yet fully understood, here we summarise the current state of knowledge. Given rising rates of GDM, now estimated to be 14% globally, and widespread FA food fortification, further research is urgently needed to elucidate the mechanisms which underpin GDM pathogenesis.

Dietary-derived vitamin B12 protects Caenorhabditis elegans from thiol-reducing agents.

BACKGROUND: One-carbon metabolism, which includes the folate and methionine cycles, involves the transfer of methyl groups which are then utilised as a part of multiple physiological processes including redox defence. During the methionine cycle, the vitamin B12-dependent enzyme methionine synthetase converts homocysteine to methionine. The enzyme S-adenosylmethionine (SAM) synthetase then uses methionine in the production of the reactive methyl carrier SAM. SAM-binding methyltransferases then utilise SAM as a cofactor to methylate proteins, small molecules, lipids, and nucleic acids. RESULTS: We describe a novel SAM methyltransferase, RIPS-1, which was the single gene identified from forward genetic screens in Caenorhabditis elegans looking for resistance to lethal concentrations of the thiol-reducing agent dithiothreitol (DTT). As well as RIPS-1 mutation, we show that in wild-type worms, DTT toxicity can be overcome by modulating vitamin B12 levels, either by using growth media and/or bacterial food that provide higher levels of vitamin B12 or by vitamin B12 supplementation. We show that active methionine synthetase is required for vitamin B12-mediated DTT resistance in wild types but is not required for resistance resulting from RIPS-1 mutation and that susceptibility to DTT is partially suppressed by methionine supplementation. A targeted RNAi modifier screen identified the mitochondrial enzyme methylmalonyl-CoA epimerase as a strong genetic enhancer of DTT resistance in a RIPS-1 mutant. We show that RIPS-1 is expressed in the intestinal and hypodermal tissues of the nematode and that treating with DTT, ß-mercaptoethanol, or hydrogen sulfide induces RIPS-1 expression. We demonstrate that RIPS-1 expression is controlled by the hypoxia-inducible factor pathway and that homologues of RIPS-1 are found in a small subset of eukaryotes and bacteria, many of which can adapt to fluctuations in environmental oxygen levels. CONCLUSIONS: This work highlights the central importance of dietary vitamin B12 in normal metabolic processes in C. elegans, defines a new role for this vitamin in countering reductive stress, and identifies RIPS-1 as a novel methyltransferase in the methionine cycle.

Profiling the Influence of Gene Variants Related to Folate-Mediated One-Carbon Metabolism on the Outcome of In Vitro Fertilization (IVF) with Donor Oocytes in Recipients Receiving Folic Acid Fortification.

Nutritional status and gene polymorphisms of one-carbon metabolism confer a well-known interaction that in pregnant women may affect embryo viability and the health of the newborn. Folate metabolism directly impacts nucleotide synthesis and methylation, which is of increasing interest in the reproductive medicine field. Studies assessing the genetic influence of folate metabolism on IVF treatments have currently been performed in women using their own oocytes. Most of these patients seeking to have a child or undergoing IVF treatments are advised to preventively intake folate supplies that restore known metabolic imbalances, but the treatments could lead to the promotion of specific enzymes in specific women, depending on their genetic variance. In the present study, we assess the influence of candidate gene variants related to folate metabolism, such as Serine Hydroxymethyltransferase 1 SHMT1 (rs1979276 and rs1979277), Betaine-Homocysteine S-Methyltransferase BHMT (rs3733890), Methionine synthase reductase MTRR (rs1801394), Methylenetetrahydrofolate reductase MTHFR (rs1801131 and rs1801133), methionine synthase MTR (rs12749581), ATP Binding Cassette Subfamily B Member 1 ABCB1 (rs1045642) and folate receptor alpha FOLR1 (rs2071010) on the success of IVF treatment performed in women being recipients of donated oocytes. The implication of such gene variants seems to have no direct impact on pregnancy consecution after IVF; however, several gene variants could influence pregnancy loss events or pregnancy maintenance, as consequence of folic acid fortification.

PhoP- and GlnR-mediated regulation of metK transcription and its impact upon S-adenosyl-methionine biosynthesis in Saccharopolyspora erythraea.

BACKGROUND: Erythromycin A (Er A) has a broad antibacterial effect and is a source of erythromycin derivatives. Methylation of erythromycin C (Er C), catalyzed by S-adenosyl-methionine (SAM)-dependent O-methyltransferase EryG, is the key final step in Er A biosynthesis. Er A biosynthesis, including EryG production, is regulated by the phosphate response factor PhoP and the nitrogen response factor GlnR. However, the regulatory effect of these proteins upon S-adenosyl-methionine synthetase (MetK) production is unknown. RESULTS: In this study, we used bioinformatics approaches to identify metK (SACE_3900), which codes for S-adenosyl-methionine synthetase (MetK). Electrophoretic mobility shift assays (EMSAs) revealed that PhoP and GlnR directly interact with the promoter of metK, and quantitative PCR (RT-qPCR) confirmed that each protein positively regulated metK transcription. Moreover, intracellular SAM was increased upon overexpression of either phoP or glnR under phosphate or nitrogen limited conditions, respectively. Finally, both the production of Er A and the transformation ratio from Er C to Er A increased upon phoP overexpression, but surprisingly, not upon glnR overexpression. CONCLUSIONS: Manipulating the phosphate and nitrogen response factors, PhoP and GlnR provides a novel strategy for increasing the yield of SAM and the production of Er A in Saccharopolyspora erythraea .

Human B12-dependent enzymes: Methionine synthase and Methylmalonyl-CoA mutase.

Humans have only two known cobalamin or B12-dependent enzymes: cytoplasmic methionine synthase and mitochondrial methylmalonyl-CoA mutase. A complex intracellular B12 trafficking pathway, comprising a multitude of chaperones, process and deliver cobalamin to the two target enzymes. Methionine synthase catalyzes the transfer of a methyl group from N5-methytetrahydrofolate to homocysteine, generating tetrahydrofolate and methionine. Cobalamin serves as an intermediate methyl group carrier and cycles between methylcobalamin and cob(I)alamin. Methylmalonyl-CoA mutase uses the 5'-deoxyadenosylcobalamin form of the cofactor and catalyzes the 1,2 rearrangement of methylmalonyl-CoA to succinyl-CoA. Two chaperones, CblA (or MMAA) and CblB (or MMAB, also known as adenosyltransferase), serve the mutase and ensure that the fidelity of the cofactor loading and unloading processes is maintained. This chapter focuses on assays for purifying and measuring the activities of methionine synthase and methylmalonyl-CoA mutase.

Causes and consequences of impaired methionine synthase activity in acquired and inherited disorders of vitamin B12 metabolism.

Methyl-Cobalamin (Cbl) derives from dietary vitamin B12 and acts as a cofactor of methionine synthase (MS) in mammals. MS encoded by MTR catalyzes the remethylation of homocysteine to generate methionine and tetrahydrofolate, which fuel methionine and cytoplasmic folate cycles, respectively. Methionine is the precursor of S-adenosyl methionine (SAM), the universal methyl donor of transmethylation reactions. Impaired MS activity results from inadequate dietary intake or malabsorption of B12 and inborn errors of Cbl metabolism (IECM). The mechanisms at the origin of the high variability of clinical presentation of impaired MS activity are classically considered as the consequence of the disruption of the folate cycle and related synthesis of purines and pyrimidines and the decreased synthesis of endogenous methionine and SAM. For one decade, data on cellular and animal models of B12 deficiency and IECM have highlighted other key pathomechanisms, including altered interactome of MS with methionine synthase reductase, MMACHC, and MMADHC, endoplasmic reticulum stress, altered cell signaling, and genomic/epigenomic dysregulations. Decreased MS activity increases catalytic protein phosphatase 2A (PP2A) and produces imbalanced phosphorylation/methylation of nucleocytoplasmic RNA binding proteins, including ELAVL1/HuR protein, with subsequent nuclear sequestration of mRNAs and dramatic alteration of gene expression, including SIRT1. Decreased SAM and SIRT1 activity induce ER stress through impaired SIRT1-deacetylation of HSF1 and hypomethylation/hyperacetylation of peroxisome proliferator-activated receptor-γ coactivator-1α (PGC1α), which deactivate nuclear receptors and lead to impaired energy metabolism and neuroplasticity. The reversibility of these pathomechanisms by SIRT1 agonists opens promising perspectives in the treatment of IECM outcomes resistant to conventional supplementation therapies.

Methionine synthase supports tumour tetrahydrofolate pools.

Mammalian cells require activated folates to generate nucleotides for growth and division. The most abundant circulating folate species is 5-methyl tetrahydrofolate (5-methyl-THF), which is used to synthesize methionine from homocysteine via the cobalamin-dependent enzyme methionine synthase (MTR). Cobalamin deficiency traps folates as 5-methyl-THF. Here, we show using isotope tracing that MTR is only a minor source of methionine in cell culture, tissues or xenografted tumours. Instead, MTR is required for cells to avoid folate trapping and assimilate 5-methyl-THF into other folate species. Under conditions of physiological extracellular folates, genetic MTR knockout in tumour cells leads to folate trapping, purine synthesis stalling, nucleotide depletion and impaired growth in cell culture and as xenografts. These defects are rescued by free folate but not one-carbon unit supplementation. Thus, MTR plays a crucial role in liberating THF for use in one-carbon metabolism.

Methionine synthase is essential for cancer cell proliferation in physiological folate environments.

Folate metabolism can be an effective target for cancer treatment. However, standard cell culture conditions utilize folic acid, a non-physiological folate source for most tissues. We find that the enzyme that couples folate and methionine metabolic cycles, methionine synthase, is required for cancer cell proliferation and tumour growth when 5-methyl tetrahydrofolate (THF), the major folate found in circulation, is the extracellular folate source. In such physiological conditions, methionine synthase incorporates 5-methyl THF into the folate cycle to maintain intracellular levels of the folates needed for nucleotide production. 5-methyl THF can sustain intracellular folate metabolism in the absence of folic acid. Therefore, cells exposed to 5-methyl THF are more resistant to methotrexate, an antifolate drug that specifically blocks folic acid incorporation into the folate cycle. Together, these data argue that the environmental folate source has a profound effect on folate metabolism, determining how both folate cycle enzymes and antifolate drugs impact proliferation.

Cuantificación, adecuación de la ingesta y fuentes alimentarias de nutrientes relacionados con el ciclo metionina-metilación (colina, betaína, folatos, vitamina B6 y vitamina B12) en mujeres embarazadas en España / Quantification, dietary intake adequacy, and food sources of nutrients involved in the methionine-methylation cycle (choline, betaine, folate, vitamin B6 and vitamin B12) in pregnant women in Spain

Objetivo: cuantificar las ingestas dietéticas de los micronutrientes implicados en el ciclo metilación-metionina (colina, betaína, folatos, vitaminas B6 y B12) en una muestra representativa de mujeres gestantes residentes en España; determinar la adecuación a las recomendaciones, y analizar sus principales fuentes alimentarias. Material y métodos: la determinación de la ingesta media se realizó a partir de los datos de consumo de los alimentos recogidos en la “Encuesta Nacional de Alimentación en población adulta, mayores y embarazadas” (ENALIA-2) (n = 133). Para el cálculo del aporte de folatos y de vitaminas B6 y B12 se emplearon los datos de composición nutricional recogidos en las “Tablas de Composición de Alimentos en España”, mientras que para la colina y la betaína, nutrientes no incluidos en las bases de datos de composición de alimentos en Europa, se empleó la “Base de Datos Nacional de Nutrientes para Referencia Estándar del Departamento de Agricultura de los Estados Unidos” (USDA). La adecuación de la ingesta se estimó de acuerdo con las recomendaciones de las principales guías españolas, europeas y estadounidenses. (AU)

Objective: a quantification of dietary intakes of the micronutrients involved in the methylation-methionine cycle (choline, betaine, folate, vitamins B6 and B12) in a representative sample of pregnant women in Spain; assessment of intake adequacy to available official recommendations; and analysis of their main food sources. Material and methods: the median intake of each micronutrient was established using food consumption data reported in the National Dietary Survey of adults, the elderly, and pregnant women (ENALIA-2) (n = 133). For folate, vitamin B6 and vitamin B12 intake, nutritional composition data from the Spanish Food Composition Tables were used, whereas for choline and betaine, which are not included in European food composition databases, the National Nutrient Database for Standard Reference of the United States Department of Agriculture (USDA) was considered. Intake adequacy was estimated in accordance with the recommendations of the main Spanish, European, and US guidelines. (AU)

Biochemical and Mutational Analysis of Radical S-Adenosyl-L-Methionine Adenosylhopane Synthase HpnH from Zymomonas mobilis Reveals that the Conserved Residue Cysteine-106 Reduces a Radical Intermediate and Determines the Stereochemistry.

Adenosylhopane is a crucial precursor of C35 hopanoids, which are believed to modulate the fluidity and permeability of bacterial cell membranes. Adenosylhopane is formed by a crosslinking reaction between diploptene and a 5'-deoxyadenosyl radical that is generated by the radical S-adenosyl-L-methionine (SAM) enzyme HpnH. We previously showed that HpnH from Streptomyces coelicolor A3(2) (ScHpnH) converts diploptene to (22R)-adenosylhopane. However, the mechanism of the stereoselective C-C bond formation was unclear. Thus, here, we performed biochemical and mutational analysis of another HpnH, from the ethanol-producing bacterium Zymomonas mobilis (ZmHpnH). Similar to ScHpnH, wild-type ZmHpnH afforded (22R)-adenosylhopane. Conserved cysteine and tyrosine residues were suggested as possible hydrogen sources to quench the putative radical reaction intermediate. A Cys106Ala mutant of ZmHpnH had one-fortieth the activity of the wild-type enzyme and yielded both (22R)- and (22S)-adenosylhopane along with some related byproducts. Radical trapping experiments with a spin-trapping agent supported the generation of a radical intermediate in the ZmHpnH-catalyzed reaction. We propose that the thiol of Cys106 stereoselectively reduces the radical intermediate generated at the C22 position by the addition of the 5'-deoxadenosyl radical to diploptene, to complete the reaction.

Vitamin B12 impacts amyloid beta-induced proteotoxicity by regulating the methionine/S-adenosylmethionine cycle.

Alzheimer's disease (AD) is a devastating neurodegenerative disorder with no effective treatment. Diet, as a modifiable risk factor for AD, could potentially be targeted to slow disease onset and progression. However, complexity of the human diet and indirect effects of the microbiome make it challenging to identify protective nutrients. Multiple factors contribute to AD pathogenesis, including amyloid beta (Aß) deposition, energy crisis, and oxidative stress. Here, we use Caenorhabditis elegans to define the impact of diet on Aß proteotoxicity. We discover that dietary vitamin B12 alleviates mitochondrial fragmentation, bioenergetic defects, and oxidative stress, delaying Aß-induced paralysis without affecting Aß accumulation. Vitamin B12 has this protective effect by acting as a cofactor for methionine synthase, impacting the methionine/S-adenosylmethionine (SAMe) cycle. Vitamin B12 supplementation of B12-deficient adult Aß animals is beneficial, demonstrating potential for vitamin B12 as a therapy to target pathogenic features of AD triggered by proteotoxic stress.