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Cultural exchange in 1954 - 1960 played an important role in the formation of Soviet-Indian cooperation. Adhering to the principles of "pancha shila", the Soviet leadership used the strategy of "soft power" to expand its own influence in Asian countries. The exchange of creative teams and individual performers laid the foundation for building cultural relations between the USSR and India in the future. Closer contacts in the period under review were established between circus, theatrical art and dance ensembles. Spectators from both countries had the opportunity to get to know the intangible cultural heritage of the USSR and India, which, in turn, formed a positive impression on the public. Without official agreements, the exchange was carried out during the period under review on the basis of mutual interest, thanks to the initiative of the USSR leadership, closer interaction was carried out, contacts were expanded in the cultural sphere. Consideration of the issue of the place of variety art in the Soviet-Indian cultural exchange makes it possible to determine the features of the process, identify problematic components, in particular, the reluctance to expand the geography of exchanges and the increase in the cost of organizing tours by the Indian leadership. Nevertheless, the Soviet side sought to use this channel to promote the Soviet way of life, deliberately investing heavily in the implementation of plans for cultural interaction with India.

Objectives: To estimate the frequency of the cystathionine beta-synthase deficiency caused by c.1105C>T mutation in Central Europe compared to Norway, and to examine the pathogenicity of the corresponding p.R369C mutant enzyme.Study Design: Mutation c.1105C>T was analyzed in 600 anonymous Czech newborn blood spots. Catalytic activity and quaternary structure of the p.R369C mutant was evaluated after expression in 2 cellular systems.Results: Population frequency of the c.1105C>T mutation was 0.005, predicting the birth prevalence of homocystinuria of 1:40000, which increased to 1:15500 in a model including 10 additional mutations. In Escherichia coli the p.R369C mutant misfolded, and its activity was severely reduced, and expression in Chinese hamster ovary cells enabled proper folding with activity decreased to 63% of the wild-type enzyme. This decreased activity was not due to impaired K(m) for both substrates but resulted from V(max) lowered to 55% of the normal cystathionine beta-synthase enzyme.Conclusions: The c.1105C>T (p.R369C) allele is common also in the Czech population. Although the p.R369C mutation impairs folding and decreases velocity of the enzymatic reaction, our data are congruent with rather mild clinical phenotype in homozygotes or compound heterozygotes carrying this mutation. [ABSTRACT FROM AUTHOR]

Background: In animals, dietary sulfur amino acid restriction (SAAR) improves metabolic health, possibly mediated by altering sulfur amino acid metabolism and enhanced anti-obesogenic processes in adipose tissue., Aim: To assess the effects of SAAR over time on the plasma and urine SAA-related metabolites (sulfurome) in humans with overweight and obesity, and explore whether such changes were associated with body weight, body fat and adipose tissue gene expression., Methods: Fifty-nine subjects were randomly allocated to SAAR (∼2 g SAA, n = 31) or a control diet (∼5.6 g SAA, n = 28) consisting of plant-based whole-foods and supplemented with capsules to titrate contents of SAA. Sulfurome metabolites in plasma and urine at baseline, 4 and 8 weeks were measured using HPLC and LC-MS/MS. mRNA-sequencing of subcutaneous white adipose tissue (scWAT) was performed to assess changes in gene expression. Data were analyzed with mixed model regression. Principal component analyses (PCA) were performed on the sulfurome data to identify potential signatures characterizing the response to SAAR., Results: SAAR led to marked decrease of the main urinary excretion product sulfate (p < 0.001) and plasma and/or 24-h urine concentrations of cystathionine, sulfite, thiosulfate, H 2 S, hypotaurine and taurine. PCA revealed a distinct metabolic signature related to decreased transsulfuration and H 2 S catabolism that predicted greater weight loss and android fat mass loss in SAAR vs. controls (all p interaction < 0.05). This signature correlated positively with scWAT expression of genes in the tricarboxylic acid cycle, electron transport and β-oxidation (FDR = 0.02)., Conclusion: SAAR leads to distinct alterations of the plasma and urine sulfurome in humans, and predicted increased loss of weight and android fat mass, and adipose tissue lipolytic gene expression in scWAT. Our data suggest that SAA are linked to obesogenic processes and that SAAR may be useful for obesity and related disorders. TRIAL IDENTIFIER: https://clinicaltrials.gov/study/NCT04701346., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Elevated total plasma homocysteine (hyperhomocysteinemia) is a marker of cardiovascular, thrombotic, and neuropsychological disease. It has multiple causes, including the common nutritional vitamin B12 or folate deficiency. However, some rare but treatable, inborn errors of metabolism (IEM) characterized by hyperhomocysteinemia can be missed due to variable presentations and the lack of awareness. The aim of this study is to identify undiagnosed IEM in adults with significantly elevated homocysteine using key existing clinical data points, then IEM specific treatment can be offered to improve outcome. We conducted a retrospective study with data mining and chart review of patients with plasma total homocysteine >30 μmol/L over a two-year period. We offer biochemical and genetic testing to patients with significant hyperhomocysteinemia without a clear explanation to diagnose IEM. We identified 22 subjects with significant hyperhomocysteinemia but no clear explanation. Subsequently, we offered genetic testing to seven patients and diagnosed one patient with classic homocystinuria due to cystathionine beta-synthase deficiency. With treatment, she lowered her plasma homocysteine and improved her health. This study stresses the importance of a thorough investigation of hyperhomocysteinemia in adults to identify rare but treatable IEM. We propose a metabolic evaluation algorithm for elevated homocysteine levels., (© 2022 Wiley Periodicals LLC.)

Decreasing the dietary intake of methionine exerts robust anti-adiposity effects in rodents but modest effects in humans. Since cysteine can be synthesized from methionine, animal diets are formulated by decreasing methionine and eliminating cysteine. Such diets exert both methionine restriction (MR) and cysteine restriction (CR), that is, sulfur amino acid restriction (SAAR). Contrarily, SAAR diets formulated for human consumption included cysteine, and thus might have exerted only MR. Epidemiological studies positively correlate body adiposity with plasma cysteine but not methionine, suggesting that CR, but not MR, is responsible for the anti-adiposity effects of SAAR. Whether this is true, and, if so, the underlying mechanisms are unknown. Using methionine- and cysteine-titrated diets, we demonstrate that the anti-adiposity effects of SAAR are due to CR. Data indicate that CR increases serinogenesis (serine biosynthesis from non-glucose substrates) by diverting substrates from glyceroneogenesis, which is essential for fatty acid reesterification and triglyceride synthesis. Molecular data suggest that CR depletes hepatic glutathione and induces Nrf2 and its downstream targets Phgdh (the serine biosynthetic enzyme) and Pepck-M. In mice, the magnitude of SAAR-induced changes in molecular markers depended on dietary fat concentration (60% fat >10% fat), sex (males > females), and age-at-onset (young > adult). Our findings are translationally relevant as we found negative and positive correlations of plasma serine and cysteine, respectively, with triglycerides and metabolic syndrome criteria in a cross-sectional epidemiological study. Controlled feeding of low-SAA, high-polyunsaturated fatty acid diets increased plasma serine in humans. Serinogenesis might be a target for treating hypertriglyceridemia., (© 2022 The Authors. Aging Cell published by Anatomical Society and John Wiley & Sons Ltd.)

Regulation of H 2 S homeostasis in humans is poorly understood. Therefore, we assessed the importance of individual enzymes in synthesis and catabolism of H 2 S by studying patients with respective genetic defects. We analyzed sulfur compounds (including bioavailable sulfide) in 37 untreated or insufficiently treated patients with seven ultrarare enzyme deficiencies and compared them to 63 controls. Surprisingly, we observed that patients with severe deficiency in cystathionine β-synthase (CBS) or cystathionine γ-lyase (CSE) - the enzymes primarily responsible for H 2 S synthesis - exhibited increased and normal levels of bioavailable sulfide, respectively. However, an approximately 21-fold increase of urinary homolanthionine in CBS deficiency strongly suggests that lacking CBS activity is compensated for by an increase in CSE-dependent H 2 S synthesis from accumulating homocysteine, which suggests a control of H 2 S homeostasis in vivo. In deficiency of sulfide:quinone oxidoreductase - the first enzyme in mitochondrial H 2 S oxidation - we found normal H 2 S concentrations in a symptomatic patient and his asymptomatic sibling, and elevated levels in an asymptomatic sibling, challenging the requirement for this enzyme in catabolizing H 2 S under physiological conditions. Patients with ethylmalonic encephalopathy and sulfite oxidase/molybdenum cofactor deficiencies exhibited massive accumulation of thiosulfate and sulfite with formation of large amounts of S-sulfocysteine and S-sulfohomocysteine, increased renal losses of sulfur compounds and concomitant strong reduction in plasma total cysteine. Our results demonstrate the value of a comprehensive assessment of sulfur compounds in severe disorders of homocysteine/cysteine metabolism and provide evidence for redundancy and compensatory mechanisms in the maintenance of H 2 S homeostasis., Competing Interests: Declaration of competing interest GS declares that he serves as CEO of Colbourne Pharmaceuticals consulting Origin Biosciences in the developments of treatments for MoCD type A, BS is investigator in clinical trials to develop a treatment for MoCD-A sponsored by Origin Biosciences Inc. The other authors do not declare any competing interests., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Cystathionine β-synthase (CBS) deficiency has a wide clinical spectrum, ranging from neurodevelopmental problems, lens dislocation and marfanoid features in early childhood to adult onset disease with predominantly thromboembolic complications. We have analysed clinical and laboratory data at the time of diagnosis in 328 patients with CBS deficiency from the E-HOD (European network and registry for Homocystinurias and methylation Defects) registry. We developed comprehensive criteria to classify patients into four groups of pyridoxine responsivity: non-responders (NR), partial, full and extreme responders (PR, FR and ER, respectively). All groups showed overlapping concentrations of plasma total homocysteine while pyridoxine responsiveness inversely correlated with plasma/serum methionine concentrations. The FR and ER groups had a later age of onset and diagnosis and a longer diagnostic delay than NR and PR patients. Lens dislocation was common in all groups except ER but the age of dislocation increased with increasing responsiveness. Developmental delay was commonest in the NR group while no ER patient had cognitive impairment. Thromboembolism was the commonest presenting feature in ER patients, whereas it was least likely at presentation in the NR group. This probably is due to the differences in ages at presentation: all groups had a similar number of thromboembolic events per 1000 patient-years. Clinical severity of CBS deficiency depends on the degree of pyridoxine responsiveness. Therefore, a standardised pyridoxine-responsiveness test in newly diagnosed patients and a critical review of previous assessments is indispensable to ensure adequate therapy and to prevent or reduce long-term complications., (© 2020 The Authors. Journal of Inherited Metabolic Disease published by John Wiley & Sons Ltd on behalf of SSIEM.)

Background and Purpose: Homocystinurias are rare genetic defects characterized by altered fluxes of sulfur compounds including homocysteine and cysteine. We explored whether the severely perturbed sulfur amino acid metabolism in patients with homocystinurias affects the metabolism of hydrogen sulfide., Experimental Approach: We studied 10 treated patients with a block in the conversion of homocysteine to cysteine due to cystathionine β-synthase deficiency (CBSD) and six treated patients with remethylation defects (RMD) and an enhanced flux of sulfur metabolites via transsulfuration. Control groups for CBSD and RMD patients consisted of 22 patients with phenylketonuria on a low-protein diet and of 12 healthy controls respectively. Plasma and urine concentrations of selected sulfur compounds were analysed by HPLC and LC-MS/MS., Key Results: Patients with CBSD exhibited plasma concentrations of monobromobimane-detected sulfide similar to appropriate controls. Urinary homolanthionine and thiosulfate in CBSD were increased significantly 1.9 and 3 times suggesting higher hydrogen sulfide synthesis by γ-cystathionase and detoxification respectively. Surprisingly, patients with RMD had significantly lower plasma sulfide levels (53 and 64% of controls) with lower sulfite concentrations, and higher taurine and thiosulfate levels suggesting enhanced cysteine oxidation and hydrogen sulfide catabolism respectively., Conclusion and Implications: The results from this study suggest that severe inherited defects in sulfur amino acid metabolism may be accompanied by only moderately perturbed hydrogen sulfide metabolism and lends support to the hypothesis that enzymes in the transsulfuration pathway may not be the major contributors to the endogenous hydrogen sulfide pool., Linked Articles: This article is part of a themed section on Chemical Biology of Reactive Sulfur Species. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.4/issuetoc., (© 2018 The British Pharmacological Society.)

Aims: The transsulfuration pathway enzymes cystathionine beta-synthase (CBS) and cystathionine gamma-lyase are thought to be the major source of hydrogen sulfide (H 2 S). In this study, we assessed the role of CBS in H 2 S biogenesis., Results: We show that despite discouraging enzyme kinetics of alternative H 2 S-producing reactions utilizing cysteine compared with the canonical condensation of serine and homocysteine, our simulations of substrate competitions at biologically relevant conditions suggest that cysteine is able to partially compete with serine on CBS, thus leading to generation of appreciable amounts of H 2 S. The leading H 2 S-producing reaction is condensation of cysteine with homocysteine, while cysteine desulfuration plays a dominant role when cysteine is more abundant than serine and homocysteine is limited. We found that the serine-to-cysteine ratio is the main determinant of CBS H 2 S productivity. Abundance of cysteine over serine, for example, in plasma, allowed for up to 43% of CBS activity being responsible for H 2 S production, while excess of serine typical for intracellular levels effectively limited such activity to less than 1.5%. CBS also produced lanthionine from serine and cysteine and a third of lanthionine coming from condensation of two cysteines contributed to the H 2 S pool., Innovation: Our study characterizes the H 2 S-producing potential of CBS under biologically relevant conditions and highlights the serine-to-cysteine ratio as the main determinant of H 2 S production by CBS in vivo., Conclusion: Our data clarify the function of CBS in H 2 S biogenesis and the role of thioethers as surrogate H 2 S markers. Antioxid. Redox Signal. 28, 311-323.

Clinical treatment of the infections caused by various staphylococcal species differ depending on the actual cause of infection. Therefore, it is necessary to develop a fast and reliable method for identification of staphylococci. Raman spectroscopy is an optical method used in multiple scientific fields. Recent studies showed that the method has a potential for use in microbiological research, too. Our work here shows a possibility to identify staphylococci by Raman spectroscopy. We present a method that enables almost 100% successful identification of 16 of the clinically most important staphylococcal species directly from bacterial colonies grown on a Mueller-Hinton agar plate. We obtained characteristic Raman spectra of 277 staphylococcal strains belonging to 16 species from a 24-hour culture of each strain grown on the Mueller-Hinton agar plate using the Raman instrument. The results show that it is possible to distinguish among the tested species using Raman spectroscopy and therefore it has a great potential for use in routine clinical diagnostics.

Background: Liver enzymes are released from hepatocytes into circulation and their activity can be measured in the blood. We examined whether the plasma activity of the liver enzyme ornithine carbamoyltransferase, determined by a novel liquid chromatography-mass spectrometry (LC-MS/MS) assay, could be utilized for the detection of OTC deficiency (OTCD), an X-linked inborn error of the urea cycle., Methods: The plasma ornithine carbamoyltransferase (OTC) activity was assayed in the reverse reaction using isotopically labeled citrulline-d4 as a substrate and by determination of the product, ornithine-d4, by LC-MS/MS analysis., Results: The plasma OTC activity in the controls was in the range of 111-658 pkat/L (n=49, median 272 pkat/L), and the activity increased linearly with serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities in patients with hepatopathy. The OTC activity was subsequently determined in 32 individuals carrying mutations in the OTC gene, and OTC/ALT and OTC/AST ratios were calculated to account for the degree of hepatopathy, which is a common finding in OTCD. The OTC/ALT ratio enabled clear differentiation of OTCD hemizygotes (n=11, range 0-69×10-6) from controls (504-3440×10-6). This ratio also enabled the detection of 11 of 12 symptomatic heterozygotes (range 38-794×10-6), while this marker did not allow for reliable differentiation of asymptomatic heterozygotes (n=9) from controls., Conclusions: LC-MS/MS assay of plasma OTC activity enabled the detection of all hemizygous and the majority of symptomatic heterozygous OTCD patients in the tested cohort. This study demonstrates that non-invasive assay of enzymes expressed predominantly in the liver could be used as an alternative approach for diagnosing inborn errors of metabolism.

Two enzymes in the transsulfuration pathway of homocysteine -cystathionine beta-synthase (CBS) and gamma-cystathionase (CTH)-use cysteine and/or homocysteine to produce the important signaling molecule hydrogen sulfide (H2S) and simultaneously the thioethers lanthionine, cystathionine or homolanthionine. In this study we explored whether impaired flux of substrates for H2S synthesis and/or deficient enzyme activities alter production of hydrogen sulfide in patients with homocystinurias. As an indirect measure of H2S synthesis we determined by LC-MS/MS concentrations of thioethers in plasma samples from 33 patients with different types of homocystinurias, in 8 patient derived fibroblast cell lines, and as reaction products of seven purified mutant CBS enzymes. Since chaperoned recombinant mutant CBS enzymes retained capacity of H2S synthesis in vitro it can be stipulated that deficient CBS activity in vivo may impair H2S production. Indeed, in patients with classical homocystinuria we observed significantly decreased cystathionine and lanthionine concentrations in plasma (46% and 74% of median control levels, respectively) and significantly lower cystathionine in fibroblasts (8% of median control concentrations) indicating that H2S production from cysteine and homocysteine may be also impaired. In contrast, the grossly elevated plasma levels of homolanthionine in CBS deficient patients (32-times elevation compared to median of controls) clearly demonstrates a simultaneous overproduction of H2S from homocysteine by CTH. In the remethylation defects the accumulation of homocysteine and the increased flux of metabolites through the transsulfuration pathway resulted in elevation of cystathionine and homolanthionine (857% and 400% of median control values, respectively) indicating a possibility of an increased biosynthesis of H2S by both CBS and CTH. This study shows clearly disturbed thioether concentrations in homocystinurias, and modeling using these data indicates that H2S synthesis may be increased in these conditions. Further studies are needed to confirm our findings and to explore the possible implications for pathophysiology of these disorders., (Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Metabolism of homocysteine and other sulfur amino acids is closely associated with metabolism of folates. In this study, we analyzed the possible role of folates and sulfur amino acids in the development of features of the metabolic syndrome in the BXH/HXB recombinant inbred strains derived from the spontaneously hypertensive rat (SHR) and Brown Norway progenitors. We mapped a quantitative trait locus for cysteine concentrations to a region of chromosome 1 that contains a cis-acting expression quantitative trait locus regulating mRNA levels of folate receptor 1 (Folr1) in the kidney. Sequence analysis revealed a deletion variant in the Folr1 promoter region of the SHR. Transfection studies demonstrated that the SHR-promoter region of Folr1 is less effective in driving luciferase reporter gene expression than the Brown Norway promoter region of Folr1. Results in the SHR.BN-chr.1 congenic strain confirmed that the SHR variant in Folr1 cosegregates with markedly reduced renal expression of Folr1 and renal folate reabsorption, decreased serum levels of folate, increased serum levels of cysteine and homocysteine, increased adiposity, ectopic fat accumulation in liver and muscle, reduced muscle insulin sensitivity, and increased blood pressure. Transgenic rescue experiments performed by expressing a Folr1 transgene in the SHR ameliorated most of the metabolic disturbances. These findings are consistent with the hypothesis that inherited variation in the expression of Folr1 in the kidney influences the development of the metabolic syndrome and constitutes a previously unrecognized genetic mechanism that may contribute to increased risk for diabetes mellitus and cardiovascular disease., (© 2015 American Heart Association, Inc.)

5,10-Methylenetetrahydrofolate reductase (MTHFR) deficiency is the most common inherited disorder of folate metabolism and causes severe hyperhomocysteinaemia. To better understand the relationship between mutation and function, we performed molecular genetic analysis of 76 MTHFR deficient patients, followed by extensive enzymatic characterization of fibroblasts from 72 of these. A deleterious mutation was detected on each of the 152 patient alleles, with one allele harboring two mutations. Sixty five different mutations (42 novel) were detected, including a common splicing mutation (c.1542G>A) found in 21 alleles. Using an enzyme assay in the physiological direction, we found residual activity (1.7%-42% of control) in 42 cell lines, of which 28 showed reduced affinity for nicotinamide adenine dinucleotide phosphate (NADPH), one reduced affinity for methylenetetrahydrofolate, five flavin adenine dinucleotide-responsiveness, and 24 abnormal kinetics of S-adenosylmethionine inhibition. Missense mutations causing virtually absent activity were found exclusively in the N-terminal catalytic domain, whereas missense mutations in the C-terminal regulatory domain caused decreased NADPH binding and disturbed inhibition by S-adenosylmethionine. Characterization of patients in this way provides a basis for improved diagnosis using expanded enzymatic criteria, increases understanding of the molecular basis of MTHFR dysfunction, and points to the possible role of cofactor or substrate in the treatment of patients with specific mutations., (© 2015 WILEY PERIODICALS, INC.)

Objective: Uric acid is the end product of purine metabolism in humans, and increased serum uric acid concentrations lead to gout. The objective of the current study was to identify factors that are independently associated with serum uric acid concentrations in a cohort of Czech control individuals., Methods: The cohort consisted of 589 healthy subjects aged 18-65 years. We studied the associations between the serum uric acid concentration and the following: (i) demographic, anthropometric and other variables previously reported to be associated with serum uric acid concentrations; (ii) the presence of metabolic syndrome and the levels of metabolic syndrome components; and (iii) selected genetic variants of the MTHFR (c.665C>T, c.1286A>C), SLC2A9 (c.844G>A, c.881G>A) and ABCG2 genes (c.421C>A). A backward model selection procedure was used to build two multiple linear regression models; in the second model, the number of metabolic syndrome criteria that were met replaced the metabolic syndrome-related variables., Results: The models had coefficients of determination of 0.59 and 0.53. The serum uric acid concentration strongly correlated with conventional determinants including male sex, and with metabolic syndrome-related variables. In the simplified second model, the serum uric acid concentration positively correlated with the number of metabolic syndrome criteria that were met, and this model retained the explanatory power of the first model. Moderate wine drinking did not increase serum uric acid concentrations, and the urate transporter ABCG2, unlike MTHFR, was a genetic determinant of serum uric acid concentrations., Conclusion: Metabolic syndrome, moderate wine drinking and the c.421C>A variant in the ABCG gene are independently associated with the serum uric acid concentration. Our model indicates that uric acid should be clinically monitored in persons with metabolic syndrome.

Background: The role of folate deficiency and associated hyperhomocysteinemia in the pathogenesis of metabolic syndrome is not fully established. In the current study, we analyzed the role of folate deficiency in pathogenesis of the metabolic syndrome in the spontaneously hypertensive rat (SHR)., Methods: Metabolic and hemodynamic traits were assessed in SHR/Ola rats fed either folate-deficient or control diet for 4 weeks starting at the age of 3 months., Results: Compared to SHRs fed a folate-replete diet, SHRs fed a folate-deficient diet showed significantly reduced serum folate (104 ± 5 vs. 11 ± 1 nmol/L, P < 0.0005) and urinary folate excretion (4.3 ± 0.6 vs. 1.2 ± 0.1 nmol/16 h, P < 0.0005) together with a near 3-fold increase in plasma total homocysteine concentration (4.5 ± 0.1 vs 13.1 ± 0.7 μmol/L, P < 0.0005), ectopic fat accumulation in liver, and impaired glucose tolerance. Folate deficiency also increased systolic blood pressure by approximately 15 mm Hg (P < 0.01). In addition, the low-folate diet was accompanied by significantly reduced activity of antioxidant enzymes and increased concentrations of lipoperoxidation products in liver, renal cortex, and heart., Conclusions: These findings demonstrate that the SHR model is susceptible to the adverse metabolic and hemodynamic effects of low dietary intake of folate. The results are consistent with the hypothesis that folate deficiency can promote oxidative stress and multiple features of the metabolic syndrome that are associated with increased risk for diabetes and cardiovascular disease.

We report on the seventh known patient with S-adenosylhomocysteine hydrolase (SAHH) deficiency presenting at birth with features resembling phosphomannomutase 2 (PMM2-CDG Ia) deficiency. Plasma methionine and total homocysteine levels were normal at 2 months and increased only after the 8th month of age. SAHH deficiency was confirmed at 4.5 years of age by showing decreased SAHH activity (11% in both erythrocytes and fibroblasts), and compound heterozygosity for a known mutation c.145C>T (p.R49C) and a novel variant c.211G>A (p.G71S) in the AHCY gene. Retrospective analysis of clinical features revealed striking similarities between SAHH deficiency and the PMM2-CDG Ia., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The role of folates as coenzymes in 1-carbon metabolism and the clinical consequences of disturbed folate metabolism are widely known. Folate status is a complex trait determined by both exogenous and endogenous factors. This study analyzed the association between 12 genetic variants and folate status in a Czech population with no folate fortification program. These 12 genetic variants were selected from 56 variant alleles found by resequencing the coding sequences and adjacent intronic regions of 6 candidate genes involved in folate metabolism or transport (FOLR1, FOLR2, FOLR3, MTHFR, PCFT, and RFC) from 29 individuals with low plasma and erythrocyte folate concentrations. Regression analyses of a cohort of 511 Czech controls not taking folate supplements revealed that only 2 variants in the MTHFR gene were associated with altered folate concentrations in plasma and/or erythrocytes. In our previous study, we observed that the common variant MTHFR c.665C > T (known as c.677C > T; p.A222V) was associated with decreased plasma folate concentrations. In the present study, we show in addition that the rare variant MTHFR c.1958C > T (p.T653M) is associated with significantly increased erythrocyte folate concentrations (P = 0.02). Multivariate regression analysis revealed that this uncommon variant, which is present in 2% of Czech control chromosomes, explains 0.9% of the total variability of erythrocyte folate concentrations; the magnitude of this effect size was comparable with that of the common MTHFR c.665C > T variant. This result indicates that the rare genetic variants may determine folate status to a similar extent as the common allelic variant.

Misfolding of mutant enzymes may play an important role in the pathogenesis of cystathionine beta-synthase (CBS) deficiency. We examined properties of a series of 27 mutant variants, which together represent 70% of known alleles observed in patients with homocystinuria due to CBS deficiency. The median amount of SDS-soluble mutant CBS polypeptides in the pellet after centrifugation of bacterial extracts was increased by 50% compared to the wild type. Moreover, mutants formed on average only 12% of tetramers and their median activity reached only 3% of the wild-type enzyme. In contrast to the wild-type CBS about half of mutants were not activated by S-adenosylmethionine. Expression at 18 degrees C substantially increased the activity of five mutants in parallel with increasing the amounts of tetramers. We further analyzed the role of solvent accessibility of mutants as a determinant of their folding and activity. Buried mutations formed on average less tetramers and exhibited 23 times lower activity than the solvent exposed mutations. In summary, our results show that topology of mutations predicts in part the behavior of mutant CBS, and that misfolding may be an important and frequent pathogenic mechanism in CBS deficiency., ((c) 2010 Wiley-Liss, Inc.)

Homozygosity or compound heterozygosity for the c.833T>C transition (p.I278 T) in the cystathionine beta-synthase (CBS) gene represents the most common cause of pyridoxine-responsive homocystinuria in Western Eurasians. However, the frequency of the pathogenic c.833C allele, as observed in healthy newborns from several European countries (q(c.833C) approximately equals 3.3 x 10(-3)), is approximately 20-fold higher than expected on the basis of the observed number of symptomatic homocystinuria patients carrying this mutation (q(c.833C) approximately equals 0.18 x 10(-3)), implying clinical underascertainment. Intriguingly, the c.833C mutation is also present in combination with a 68-bp insertion, c.[833C; 844&#95;845ins68], in a substantial proportion of chromosomes from nonhomocystinuric individuals worldwide. We have sought to study the relationship between the pathogenic and nonpathogenic c.833C-bearing chromosomes and to determine whether the pathogenic c.[833C; -] chromosomes are identical-by-descent or instead arose by recurrent mutation. Initial haplotype analysis of 780 randomly selected Czech and sub-Saharan African wild-type chromosomes, employing 12 intragenic markers, revealed 29 distinct CBS haplotypes, of which 10 carried the c.[833C; 844&#95;845ins68] combination; none carried an isolated c.833C or c.844&#95;845ins68 mutation. Subsequent examination of 69 pathogenic c.[833C; -] chromosomes, derived from homocystinuria patients of predominantly European origin, disclosed three unrelated haplotypes that differed from their wild-type counterparts by virtue of the presence of c.833C, thereby indicating that c.833T>C transition has occurred repeatedly and independently in the past. Since c.833T does not reside within an obvious mutational hotspot, we surmise that the three pathogenic and comparatively prevalent c.[833C; -] chromosomes may have originated by recurrent gene conversion employing the common nonpathogenic c.[833C; 844&#95;845ins68] chromosomes as templates.

Homocystinuria due to cystathionine beta-synthase (CBS) deficiency is an inherited disorder of homocysteine transsulfuration, which manifests by neurological, vascular and connective tissue involvement. So far, 130 pathogenic mutations have been recognized in the CBS gene. We examined 10 independent alleles in Polish patients suffering from CBS deficiency, and we detected four already described mutations (c.1224-2A>C, c.684C>A, c.833T>C, and c.442G>A) and two novel mutations (c.429C>G and c.1039+1G>T). The pathogenicity of the novel mutations was demonstrated by expression in E.coli. This is the first published communication on mutations leading to CBS deficiency in Poland., (Copyright 2004 Wiley-Liss, Inc.)

During the past 20 years, cystathionine beta-synthase (CBS) deficiency has been detected in the former Czechoslovakia with a calculated frequency of 1:349,000. The clinical manifestation was typical of homocystinuria, and about half of the 21 patients were not responsive to pyridoxine. Twelve distinct mutations were detected in 30 independent homocystinuric alleles. One half of the alleles carried either the c.833 T-->C or the IVS11-2A-->C mutation; the remaining alleles contained private mutations. The abundance of five mutant mRNAs with premature stop codons was analyzed by PCR-RFLP. Two mRNAs, c.828&#95;931ins104 (IVS7+1G-->A) and c.1226 G-->A, were severely reduced in the cytoplasm as a result of nonsense-mediated decay. In contrast, the other three mRNAs-c.19&#95;20insC, c.28&#95;29delG, and c.210&#95;235del26 (IVS1-1G-->C)-were stable. Native western blot analysis of 14 mutant fibroblast lines showed a paucity of CBS antigen, which was detectable only in aggregates. Five mutations-A114V (c.341C-->T), A155T (c.463G-->A), E176K (c.526G-->A), I278T (c.833T-->C), and W409&#95;G453del (IVS11-2A-->C)-were expressed in Escherichia coli. All five mutant proteins formed substantially more aggregates than did the wild-type CBS, and no aggregates contained heme. These data suggest that abnormal folding, impaired heme binding, and aggregation of mutant CBS polypeptides may be common pathogenic mechanisms in CBS deficiency.

Cystathionine beta-synthase [CBS; l-serine hydro-lyase (adding homocysteine), EC 4.2.1.22] catalyzes the first committed step of transsulfuration and is the enzyme deficient in classical homocystinuria. In this report, we describe the molecular cloning and the complete nucleotide sequence of the human CBS gene. We report a total of 28,046 nucleotides of sequence, which, in addition to the CBS gene, contains approximately 5 kb of the 5' flanking region. The human CBS gene contains 23 exons ranging from 42 to 209 bp. The 5' UTR is formed by 1 of 5 alternatively used exons and 1 invariably present exon, while the 3' UTR is encoded by exons 16 and 17. We also describe the identification of two alternatively used promoter regions that are GC rich (approximately 80%) and contain numerous putative binding sites for Sp1, Ap1, Ap2, and c-myb, but lack the classical TATA box. The CBS locus contains an unusually high number of Alu repeats, which may predispose this gene to deleterious rearrangements. Additionally, we report on a number of DNA sequence repeats that are polymorphic in North American and European Caucasians., (Copyright 1998 Academic Press.)