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Background: Gutka chewing is the most common deleterious oral habit prevalent in the geographical distribution of the Indian subcontinent. Gutka leads to the production of numerous free radicals, which causes oxidative stress in regional oral tissues. Oxidative stress brings about the oxidation of guanine bases of DNA that generates 8-OHdG as its main byproduct. The presence of 8-OHdG can be evaluated not only in tissue but also in saliva, blood and urine. The availability of 8-OHdG in these samples is quite documented. In addition, a comparative assay of 8-ohdg DNA damage marker in multiple samples is yet to be done. Material and Methodology: A sample size of 60 was divided into two groups, i.e., gutka consumers without any lesion and gutka consumers with OSMF. Ten samples each of saliva, serum and urine were collected from these two groups and healthy controls. Samples were centrifuged at 1000 RPM at 2-8°C for 15-20 minutes. A volume of 1.5 ml resultant supernatant was pipetted out in labelled Eppendorf tubes and stored at -80°C. The ELISA test was performed to measure the concentration of 8-OHdG protein in different samples at 450 nm after adding stop solution in 96-well microplate. Results: 8-OHdG concentration was found to be highest in saliva followed by urine and serum. 8-OHdG concentration in serum was significantly less than that in saliva and urine (P-value <0.05). Intergroup difference in concentration of 8-OHdG of urine, saliva and serum was significant (P-value <0.05). Post hoc analysis revealed that concentration of 8-OHdG in saliva and urine was non-significantly different (P-value >0.05). Conclusion: Saliva appears to be the most appropriate sample type as compared to serum and urine for the evaluation of 8-OHdG in OSMF subjects.
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The whole field of synthetic biology (SynBio) is only about 20 years old, and plant SynBio is younger still. Nevertheless, within that short time, SynBio in general has drawn more scientific, philosophical, government, and private-sector interest than anything in biology since the recombinant DNA revolution. Plant SynBio, in particular, is now drawing more and more interest in relation to plants' potential to help solve planetary problems such as carbon capture and storage and replacing fossil fuels and feedstocks. As plant SynBio is so young and so fast-developing, we felt it was too soon to try to analyze its history. Instead, we set out to capture the essence of plant SynBio's origins and early development through interviews with 8 of the field's founders, representing 5 countries and 3 continents. We then distilled these founders' personal recollections and reflections into this review, centering the narrative on timelines for pivotal events, articles, funding programs, and quoting from interviews. We have archived the interview recordings and documented timeline entries. This work provides a resource for future historical scholarship.
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Plantas , Biología Sintética , Biología Sintética/métodos , Historia del Siglo XX , Historia del Siglo XXI , Plantas/genéticaRESUMEN
Background: The purpose of this in vitro study was to assess the effect of Tooth Mousse Plus, Remin Pro, and Fluor Protector Gel on enamel erosion, measuring mean weight loss after exposure to a demineralizing agent. Materials and methods: A total of 60 sound-extracted permanent incisors were sectioned and enamel specimens were randomly distributed to different groups. The initial weight of all specimens was registered. The samples were randomly divided into four groups (n = 30). Group I specimens were treated with tap water (control). Groups II, III, and IV were treated with Tooth Mousse, Remin Pro, and Fluor Protector Gel application. After that, specimens were placed all together in a plastic container with 6 mL of a soft drink and immersed for 8 minutes at room temperature, dried, and weighed. Specimens were weighed after each immersion period and mean weight loss was calculated. The data was analyzed for probability distribution using the Kolmogorov-Smirnov test. The intergroup comparison was done using a one-way analysis of variance (ANOVA) followed by post hoc analysis. Results: According to pairwise comparisons in post hoc analysis, the weight of specimens at baseline was significantly greater than the weight of specimens on day 12. The difference in the mean weight of the specimen from baseline to day 12 was 2.833 mg for group I, 2.367 mg for group II, 1.467 mg for group III, and 2.133 mg for group IV. Conclusion: Tooth Mousse Plus, Remin Pro, and Fluor Protector Gel have no significant effect on dental erosion. How to cite this article: Shukla K, Saxena A, Joshi J, et al. A Comparative Study of the Effect of Tooth Mousse Plus Remin Pro and Fluor Protector Gel on Enamel Erosion: An In Vitro Study. Int J Clin Pediatr Dent 2023;16(S-1):S57-S62.
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Hepatocellular carcinoma (HCC) is an aggressive liver malignancy that is difficult to treat with no approved biomarker based targeted therapies. FGF19-FGFR4 signaling blockade has been recently identified as a promising avenue for treatment of a subset of HCC patients. Using HCC relevant xenograft and PDX models, we show that Lenvatinib, an approved multi-kinase inhibitor, strongly enhanced the efficacy of FGFR4 inhibitor H3B-6527. This enhanced combination effect is not due to enhanced FGFR4 inhibition and it is likely due to cell non-autonomous VEGFR activity of Lenvatinib. This cell non-autonomous mode of action was further supported by strong in vivo combination efficacy with the mouse specific VEGFR2 antibody, DC101, which cannot cell-autonomously inhibit pathways in human xenografts. Mechanistic studies showed that the combination resulted in enhanced efficacy through increased anti-angiogenic and anti-tumorigenic activities. Overall, our results indicate that this combination can be a highly effective treatment option for FGF19 driven HCC patients, and provide preclinical validation of a combination that can be readily tested in the clinical setting.
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Continuous directed evolution of enzymes and other proteins in microbial hosts is capable of outperforming classical directed evolution by executing hypermutation and selection concurrently in vivo, at scale, with minimal manual input. Provided that a target enzyme's activity can be coupled to growth of the host cells, the activity can be improved simply by selecting for growth. Like all directed evolution, the continuous version requires no prior mechanistic knowledge of the target. Continuous directed evolution is thus a powerful way to modify plant or non-plant enzymes for use in plant metabolic research and engineering. Here, we first describe the basic features of the yeast (Saccharomyces cerevisiae) OrthoRep system for continuous directed evolution and compare it briefly with other systems. We then give a step-by-step account of three ways in which OrthoRep can be deployed to evolve primary metabolic enzymes, using a THI4 thiazole synthase as an example and illustrating the mutational outcomes obtained. We close by outlining applications of OrthoRep that serve growing demands (i) to change the characteristics of plant enzymes destined for return to plants, and (ii) to adapt ("plantize") enzymes from prokaryotes-especially exotic prokaryotes-to function well in mild, plant-like conditions.
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Evolución Molecular Dirigida/métodos , Enzimas/genética , Fitomejoramiento/métodos , Proteínas de Plantas/genética , Saccharomyces cerevisiae/genéticaRESUMEN
Abiotic stresses reduce crop growth and yield in part by disrupting metabolic homeostasis and triggering responses that change the metabolome. Experiments designed to understand the mechanisms underlying these metabolomic responses have usually not used agriculturally relevant stress regimes. We therefore subjected maize plants to drought, salt, or heat stresses that mimic field conditions and analyzed leaf responses at metabolome and transcriptome levels. Shared features of stress metabolomes included synthesis of raffinose, a compatible solute implicated in tolerance to dehydration. In addition, a marked accumulation of amino acids including proline, arginine, and γ-aminobutyrate combined with depletion of key glycolysis and tricarboxylic acid cycle intermediates indicated a shift in balance of carbon and nitrogen metabolism in stressed leaves. Involvement of the γ-aminobutyrate shunt in this process is consistent with its previously proposed role as a workaround for stress-induced thiamin-deficiency. Although convergent metabolome shifts were correlated with gene expression changes in affected pathways, patterns of differential gene regulation induced by the three stresses indicated distinct signaling mechanisms highlighting the plasticity of plant metabolic responses to abiotic stress.
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The thiamin-requiring mutants of Arabidopsis have a storied history as a foundational model for biochemical genetics in plants and have illuminated the central role of thiamin in metabolism. Recent integrative genetic and biochemical analyses of thiamin biosynthesis and utilization imply that leaf metabolism normally operates close to thiamin-limiting conditions. Thus, the mechanisms that allocate thiamin-diphosphate (ThDP) cofactor among the diverse thiamin-dependent enzymes localized in plastids, mitochondria, peroxisomes, and the cytosol comprise an intricate thiamin economy. Here, we show that the classical thiamin-requiring 3 (th3) mutant is a point mutation in plastid localized 5-deoxyxylulose synthase 1 (DXS1), a key regulated enzyme in the methylerythritol 4-phosphate (MEP) isoprene biosynthesis pathway. Substitution of a lysine for a highly conserved glutamate residue (E323) located at the subunit interface of the homodimeric enzyme conditions a hypomorphic phenotype that can be rescued by supplying low concentrations of thiamin in the medium. Analysis of leaf thiamin vitamers showed that supplementing the medium with thiamin increased total ThDP content in both wild type and th3 mutant plants, supporting a hypothesis that the mutant DXS1 enzyme has a reduced affinity for the ThDP cofactor. An unexpected upregulation of a suite of biotic-stress-response genes associated with accumulation of downstream MEP intermediate MEcPP suggests that th3 causes mis-regulation of DXS1 activity in thiamin-supplemented plants. Overall, these results highlight that the central role of ThDP availability in regulation of DXS1 activity and flux through the MEP pathway.
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Plant and fungal THI4 thiazole synthases produce the thiamin thiazole moiety in aerobic conditions via a single-turnover suicide reaction that uses an active-site Cys residue as sulfur donor. Multiple-turnover (i.e. catalytic) THI4s lacking an active-site Cys (non-Cys THI4s) that use sulfide as sulfur donor have been biochemically characterized -- but only from archaeal methanogens that are anaerobic, O2-sensitive hyperthermophiles from sulfide-rich habitats. These THI4s prefer iron as cofactor. A survey of prokaryote genomes uncovered non-Cys THI4s in aerobic mesophiles from sulfide-poor habitats, suggesting that multiple-turnover THI4 operation is possible in aerobic, mild, low-sulfide conditions. This was confirmed by testing 23 representative non-Cys THI4s for complementation of an Escherichia coli ΔthiG thiazole auxotroph in aerobic conditions. Sixteen were clearly active, and more so when intracellular sulfide level was raised by supplying Cys, demonstrating catalytic function in the presence of O2 at mild temperatures and indicating use of sulfide or a sulfide metabolite as sulfur donor. Comparative genomic evidence linked non-Cys THI4s with proteins from families that bind, transport, or metabolize cobalt or other heavy metals. The crystal structure of the aerotolerant bacterial Thermovibrio ammonificans THI4 was determined to probe the molecular basis of aerotolerance. The structure suggested no large deviations compared with the structures of THI4s from O2-sensitive methanogens, but is consistent with an alternative catalytic metal. Together with complementation data, use of cobalt rather than iron was supported. We conclude that catalytic THI4s can indeed operate aerobically and that the metal cofactor inserted is a likely natural determinant of aerotolerance.
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Archaea/enzimología , Proteínas Arqueales/química , Proteínas Arqueales/metabolismo , Bacterias/enzimología , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/enzimología , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/enzimología , Tiamina/biosíntesis , Proteínas Arqueales/genética , Biocatálisis , Dominio Catalítico , Cobalto/metabolismo , Cristalización , Cisteína/metabolismo , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Genómica/métodos , Hierro/metabolismo , Microorganismos Modificados Genéticamente , Oxígeno/metabolismo , Saccharomyces cerevisiae/genética , Sulfuros/metabolismo , Azufre/metabolismoRESUMEN
Metabolic engineering uses enzymes as parts to build biosystems for specified tasks. Although a part's working life and failure modes are key engineering performance indicators, this is not yet so in metabolic engineering because it is not known how long enzymes remain functional in vivo or whether cumulative deterioration (wear-out), sudden random failure, or other causes drive replacement. Consequently, enzymes cannot be engineered to extend life and cut the high energy costs of replacement. Guided by catalyst engineering, we adopted catalytic cycles until replacement (CCR) as a metric for enzyme functional life span in vivo. CCR is the number of catalytic cycles that an enzyme mediates in vivo before failure or replacement, i.e., metabolic flux rate/protein turnover rate. We used estimated fluxes and measured protein turnover rates to calculate CCRs for â¼100-200 enzymes each from Lactococcus lactis, yeast, and Arabidopsis CCRs in these organisms had similar ranges (<103 to >107) but different median values (3-4 × 104 in L. lactis and yeast versus 4 × 105 in Arabidopsis). In all organisms, enzymes whose substrates, products, or mechanisms can attack reactive amino acid residues had significantly lower median CCR values than other enzymes. Taken with literature on mechanism-based inactivation, the latter finding supports the proposal that 1) random active-site damage by reaction chemistry is an important cause of enzyme failure, and 2) reactive noncatalytic residues in the active-site region are likely contributors to damage susceptibility. Enzyme engineering to raise CCRs and lower replacement costs may thus be both beneficial and feasible.
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Arabidopsis/enzimología , Biocatálisis , Enzimas/química , Lactococcus lactis/enzimología , Ingeniería Metabólica , Saccharomyces cerevisiae/enzimologíaRESUMEN
Melanotic neuroectodermal tumor is a rare benign congenital neoplasm involving the head-and-neck region found in infants <1 year of age. The lesion most commonly affects the maxilla. We report the case of a 4-month-old male child with a tumor involving the anterior region of the maxilla along with a brief review of literature, differential diagnosis, and treatment.
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Tumor Neuroectodérmico Melanótico , Niño , Diagnóstico Diferencial , Humanos , Lactante , Masculino , Maxilar , Tumor Neuroectodérmico Melanótico/diagnóstico , Tumor Neuroectodérmico Melanótico/diagnóstico por imagen , Tumor Neuroectodérmico Melanótico/cirugíaRESUMEN
Plant evolution has produced enzymes that may not be optimal for maximizing yield and quality in today's agricultural environments and plant biotechnology applications. By improving enzyme performance, it should be possible to alleviate constraints on yield and quality currently imposed by kinetic properties or enzyme instability. Enzymes can be optimized more quickly than naturally possible by applying directed evolution, which entails mutating a target gene in vitro and screening or selecting the mutated gene products for the desired characteristics. Continuous directed evolution is a more efficient and scalable version that accomplishes the mutagenesis and selection steps simultaneously in vivo via error-prone replication of the target gene and coupling of the host cell's growth rate to the target gene's function. However, published continuous systems require custom plasmid assembly, and convenient multipurpose platforms are not available. We discuss two systems suitable for continuous directed evolution of enzymes, OrthoRep in Saccharomyces cerevisiae and EvolvR in Escherichia coli, and our pilot efforts to adapt each system for high-throughput plant enzyme engineering. To test our modified systems, we used the thiamin synthesis enzyme THI4, previously identified as a prime candidate for improvement. Our adapted OrthoRep system shows promise for efficient plant enzyme engineering.
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Like fungi and some prokaryotes, plants use a thiazole synthase (THI4) to make the thiazole precursor of thiamin. Fungal THI4s are suicide enzymes that destroy an essential active-site Cys residue to obtain the sulfur atom needed for thiazole formation. In contrast, certain prokaryotic THI4s have no active-site Cys, use sulfide as sulfur donor, and are truly catalytic. The presence of a conserved active-site Cys in plant THI4s and other indirect evidence implies that they are suicidal. To confirm this, we complemented the Arabidopsistz-1 mutant, which lacks THI4 activity, with a His-tagged Arabidopsis THI4 construct. LC-MS analysis of tryptic peptides of the THI4 extracted from leaves showed that the active-site Cys was predominantly in desulfurated form, consistent with THI4 having a suicide mechanism in planta. Unexpectedly, transcriptome data mining and deep proteome profiling showed that barley, wheat, and oat have both a widely expressed canonical THI4 with an active-site Cys, and a THI4-like paralog (non-Cys THI4) that has no active-site Cys and is the major type of THI4 in developing grains. Transcriptomic evidence also indicated that barley, wheat, and oat grains synthesize thiamin de novo, implying that their non-Cys THI4s synthesize thiazole. Structure modeling supported this inference, as did demonstration that non-Cys THI4s have significant capacity to complement thiazole auxotrophy in Escherichia coli. There is thus a prima facie case that non-Cys cereal THI4s, like their prokaryotic counterparts, are catalytic thiazole synthases. Bioenergetic calculations show that, relative to suicide THI4s, such enzymes could save substantial energy during the grain-filling period.
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Proteínas de Arabidopsis , Arabidopsis , Ligasas , Modelos Moleculares , Plantas Modificadas Genéticamente , Tiamina , Tiazoles/metabolismo , Arabidopsis/enzimología , Arabidopsis/genética , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Catálisis , Biología Computacional , Escherichia coli/enzimología , Escherichia coli/genética , Prueba de Complementación Genética , Ligasas/química , Ligasas/genética , Ligasas/metabolismo , Plantas Modificadas Genéticamente/enzimología , Plantas Modificadas Genéticamente/genética , Dominios Proteicos , Tiamina/biosíntesis , Tiamina/genéticaRESUMEN
Quercetin and coumarin, two naturally occurring phytochemicals of plant origin, are known to regulate hyperglycemia and oxidative stress. The present study was designed to evaluate the inhibitory activity of quercetin and coumarin on dipeptidyl peptidase-IV (DPP-IV) and their antioxidant potential. DPP-IV inhibition assays were performed, and evaluated IC50 values of diprotin A, quercetin, coumarin, and sitagliptin were found to be 0.653, 4.02, 54.83, and 5.49 nmol/mL, respectively. Furthermore, in silico studies such as the drug-likeliness and docking efficiency of quercetin and coumarin to the DPP-IV protein were performed; the ex vivo antiperoxidative potential of quercetin and coumarin were also evaluated. The results of the present study showed that the DPP-IV inhibitory potential of quercetin was slightly higher than that of sitagliptin. Virtual docking revealed the tight binding of quercetin with DPP-IV protein. Quercetin and coumarin reduced oxidative stress in vitro and ex vivo systems. We report for the first time that both compounds inhibited the DPP-IV along with antioxidant activity and thus may be use as function food ingredients in the prevention of diabetes.
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Antioxidantes/farmacología , Cumarinas/farmacología , Inhibidores de la Dipeptidil-Peptidasa IV/farmacología , Quercetina/farmacología , Animales , Simulación por Computador , Dipeptidil Peptidasa 4/metabolismo , Eritrocitos/metabolismo , Hemólisis , Enlace de Hidrógeno , Técnicas In Vitro , Concentración 50 Inhibidora , Ligandos , Peroxidación de Lípido , Hígado/metabolismo , Masculino , Conformación Molecular , Simulación del Acoplamiento Molecular , Estrés Oxidativo , Ratas , Ratas WistarRESUMEN
The suboptimal content of sulfur-containing amino acids methionine and cysteine prevents common bean (Phaseolus vulgaris) from being an excellent source of protein. Nutritional improvements to this significant crop require a better understanding of the biosynthesis of sulfur-containing compounds including the nonproteogenic amino acid S-methylcysteine and the dipeptide γ-glutamyl-S-methylcysteine, which accumulate in seed. In this study, seeds were incubated with isotopically labelled serine, cysteine or methionine and analyzed by reverse phase chromatography-high resolution mass spectrometry to track stable isotopes as they progressed through the sulfur metabolome. We determined that serine and methionine are the sole precursors of free S-methylcysteine in developing seeds, indicating that this compound is likely to be synthesized through the condensation of O-acetylserine and methanethiol. BSAS4;1, a cytosolic ß-substituted alanine synthase preferentially expressed in developing seeds, catalyzed the formation of S-methylcysteine in vitro. A higher flux of labelled serine or cysteine was observed in a sequential pathway involving γ-glutamyl-cysteine, homoglutathione and S-methylhomoglutathione, a likely precursor to γ-glutamyl-S-methylcysteine. Preferential incorporation of serine over cysteine supports a subcellular compartmentation of this pathway, likely to be in the chloroplast. The origin of the methyl group in S-methylhomoglutathione was traced to methionine. There was substantial incorporation of carbons from methionine into the ß-alanine portion of homoglutathione and S-methylhomoglutathione, suggesting the breakdown of methionine by methionine γ-lyase and conversion of α-ketobutyrate to ß-alanine via propanoate metabolism. These findings delineate the biosynthetic pathways of the sulfur metabolome of common bean and provide an insight that will aid future efforts to improve nutritional quality.
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Isótopos de Carbono/metabolismo , Cisteína/análogos & derivados , Espectrometría de Masas/métodos , Isótopos de Nitrógeno/metabolismo , Phaseolus/metabolismo , Semillas/metabolismo , Vías Biosintéticas , Liasas de Carbono-Azufre/metabolismo , Cromatografía de Fase Inversa/métodos , Cisteína/biosíntesis , Cisteína/metabolismo , Cisteína Sintasa/metabolismo , Metabolómica/métodos , Metionina/metabolismo , Serina/metabolismo , Azufre/metabolismoRESUMEN
Some grain legumes store sulfur in the form of non-protein amino acids in seed. γ-Glutamyl-S-methylcysteine is found in Phaseolus and several Vigna species. γ-Glutamyl-S-ethenylcysteine, an antinutritional compound, is present in Vicia narbonensis. In P. vulgaris, free S-methylcysteine levels are higher at early stages of seed development followed by a decline. γ-Glutamyl-S-methylcysteine accumulates later, in two phases, with a lag during reserve accumulation. The concentration of total S-methylcysteine, quantified after acid hydrolysis, is positively regulated by sulfate nutrition. The levels of both γ-glutamyl-S-methylcysteine and γ-glutamyl-S-ethenylcysteine are modulated in response to changes in seed protein composition. A model is proposed whereby the majority of γ-glutamyl-S-methylcysteine in P. vulgaris is synthesized via the intermediate S-methylhomoglutathione. Knowledge of the biosynthesis of non-protein sulfur amino acids is required for metabolic engineering approaches, in conjunction with manipulation of the protein sink, to increase the concentration of nutritionally essential methionine and cysteine. This would improve protein quality of some important legume crops.
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Aminoácidos Sulfúricos/biosíntesis , Phaseolus/metabolismo , Azufre/metabolismo , Vigna/metabolismo , Aminoácidos Sulfúricos/análisis , Vías Biosintéticas , Phaseolus/química , Phaseolus/genética , Semillas/química , Semillas/metabolismo , Vigna/química , Vigna/genéticaRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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The common bean (Phaseolus vulgaris) constitutes an excellent source of vegetable dietary protein. However, there are sub-optimal levels of the essential amino acids, methionine and cysteine. On the other hand, P. vulgaris accumulates large amounts of the γ-glutamyl dipeptide of S-methylcysteine, and lower levels of free S-methylcysteine and S-methylhomoglutathione. Past results suggest two distinct metabolite pools. Free S-methylcysteine levels are high at the beginning of seed development and decline at mid-maturation, while there is a biphasic accumulation of γ-glutamyl-S-methylcysteine, at early cotyledon and maturation stages. A possible model involves the formation of S-methylcysteine by cysteine synthase from O-acetylserine and methanethiol, whereas the majority of γ-glutamyl-S-methylcysteine may arise from S-methylhomoglutathione. Metabolite profiling during development and in genotypes differing in total S-methylcysteine accumulation showed that γ-glutamyl-S-methylcysteine accounts for most of the total S-methylcysteine in mature seed. Profiling of transcripts for candidate biosynthetic genes indicated that BSAS4;1 expression is correlated with both the developmental timing and levels of free S-methylcysteine accumulated, while homoglutathione synthetase (hGS) expression was correlated with the levels of γ-glutamyl-S-methylcysteine. Analysis of S-methylated phytochelatins by liquid chromatography and high resolution tandem mass spectrometry revealed only small amounts of homophytochelatin-2 with a single S-methylcysteine. The mitochondrial localization of phytochelatin synthase 2-predominant in seed, determined by confocal microscopy of a fusion with the yellow fluorescent protein-and its spatial separation from S-methylhomoglutathione may explain the lack of significant accumulation of S-methylated phytochelatins.
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Plants synthesize the thiazole precursor of thiamin (cThz-P) via THIAMIN4 (THI4), a suicide enzyme that mediates one reaction cycle and must then be degraded and resynthesized. It has been estimated that this THI4 turnover consumes 2% to 12% of the maintenance energy budget and that installing an energy-efficient alternative pathway could substantially increase crop yield potential. Available data point to two natural alternatives to the suicidal THI4 pathway: (i) nonsuicidal prokaryotic THI4s that lack the active-site Cys residue on which suicide activity depends, and (ii) an uncharacterized thiazole synthesis pathway in flowers of the tropical arum lily Caladium bicolor that enables production and emission of large amounts of the cThz-P analog 4-methyl-5-vinylthiazole (MVT). We used functional complementation of an Escherichia coli ΔthiG strain to identify a nonsuicidal bacterial THI4 (from Thermovibrio ammonificans) that can function in conditions like those in plant cells. We explored whether C. bicolor synthesizes MVT de novo via a novel route, via a suicidal or a nonsuicidal THI4, or by catabolizing thiamin. Analysis of developmental changes in MVT emission, extractable MVT, thiamin level, and THI4 expression indicated that C. bicolor flowers make MVT de novo via a massively expressed THI4 and that thiamin is not involved. Functional complementation tests indicated that C. bicolor THI4, which has the active-site Cys needed to operate suicidally, may be capable of suicidal and - in hypoxic conditions - nonsuicidal operation. T. ammonificans and C. bicolor THI4s are thus candidate parts for rational redesign or directed evolution of efficient, nonsuicidal THI4s for use in crop improvement.