The methionine sulphoxide reductase activity of the yeast dimethyl sulphoxide reductase system

Abstract Glycyl- l -methionine sulphoxide and N-acetyl- l -methionine sulphoxide were less effective inhibitors of the dimethyl sulphoxide (DMSO) reductase activity of Saccharomyces cerevisiae NCYC240 than was l -methionine (±)-sulphoxide. Methionine sulphoxide reductase and DMSO reductase activities from crude extracts co-purified over five purification steps and the two activities eluted from columns in exactly the same fractions. Both activities were sensitive to the same inhibitors. It was concluded that: (1) the DMSO reductase activity of S. cerevisiae is a property of a methionine sulphoxide reductase different from that of Black et al. [J. Biol. Chem. 235 (1960) 2910–2916], and (2) free methionine sulphoxide rather than peptidyl methionine sulphoxide is probably the enzyme's true substrate.     

Chromatographic separation of a thymus fraction active on lipidic metabolism

Three protein fractions have been obtained from a thymus homogenate of young calf, by a multistep sequence of procedures.Every fraction was administered by intraperitoneal means to a distinct group of rats previously fed, for 21 days, on a diet with high content of dietary fat and cholesterol.The serum values of cholesterol and triglycerids in these animals show a considerable decrease, if compared with the data obtained from control animals, especially with the third chromatographic fraction.     

Chromatographic separation of synthetic nopaline and isonopaline and their absolute configuration

Synthetic nopaline and isonopaline were separated preparatively by anion exchange chromatography and their configurations were ascertained by an enzymi     

Origin and evolution of the protein-repairing enzymes methionine sulphoxide reductases

Abstract The majority of extant life forms thrive in an O(2)-rich environment, which unavoidably induces the production of reactive oxygen species (ROS) during cellular activities. ROS readily oxidize methionine (Met) residues in proteins/peptides to form methionine sulphoxide [Met(O)] that can lead to impaired protein function. Two methionine sulphoxide reductases, MsrA and MsrB, catalyse the reduction of the S and R epimers, respectively, of Met(O) in proteins to Met. The Msr system has two known functions in protecting cells against oxidative damage. The first is to repair proteins that have lost activity due to Met oxidation and the second is to function as part of a scavenger system to remove ROS through the reversible oxidation/reduction of Met residues in proteins. Bacterial, plant and animal cells lacking MsrA are known to be more sensitive to oxidative stress. The Msr system is considered an important cellular defence mechanism to protect against oxidative stress and may be involved in ageing/senescence. MsrA is present in all known eukaryotes and eubacteria and a majority of archaea, reflecting its essential role in cellular life. MsrB is found in all eukaryotes and the majority of eubacteria and archaea but is absent in some eubacteria and archaea, which may imply a less important role of MsrB compared to MsrA. MsrA and MsrB share no sequence or structure homology, and therefore probably emerged as a result of independent evolutionary events. The fact that some archaea lack msr genes raises the question of how these archaea cope with oxidative damage to proteins and consequently of the significance of msr evolution in oxic eukaryotes dealing with oxidative stress. Our best hypothesis is that the presence of ROS-destroying enzymes such as peroxiredoxins and a lower dissolved O(2) concentration in those msr-lacking organisms grown at high temperatures might account for the successful survival of these organisms under oxidative stress.     

Trace analysis of albendazole and its sulphoxide and sulphone metabolites in milk by liquid chromatography

Analytical methodology for determination of albendazole and its sulphoxide and sulphone metabolites in milk at levels down to 2–5 ng/ml has been developed. Extraction was carried out with ethyl acetate under alkaline conditions, and extracts were analyzed on a silica-based C₁₈ column in the presence of positively-charged pairing ions. Accuracy data showed overall recoveries ranged from 78.4% to 100%, whereas precision data, based on within- and between-day variation, suggested overall precision values better than 4.9%. The method was successfully applied to determine residues in milk of a dairy cow orally given albendazole.     

Characterization of a strawberry late-expressed and fruit-specific peptide methionine sulphoxide reductase

We have cloned and characterized a cDNA clone, called Fapmsr , coding for a putative peptide methionine sulphoxide [Met(O)] reductase (PMSR, EC 1.8.4.6) from strawberry fruits ( Fragaria x ananassa ). This gene is involved in the repair of inactive peptides and proteins caused for the oxidation of methionine residues to Met(O). Expression of the Fapmsr was only detected in the receptacles of red mature fruits and not in young or immature fruits nor in other plant tissues such as flowers, leaves, runners, roots or achenes. Expression of the Fapmsr gene was activated in green immature fruits when achenes were removed from receptacles, and this was prevented by the application of exogenous auxins such as naphthaleneacetic acid. The enzyme produced and purified by cloning the strawberry cDNA in frame with the C-terminal sequence of the glutathione S-transferase gene can reduce free Met(O) to methionine as analysed by reverse phase high performance liquid chromatography. We have also set up a PMSR protection assay that demonstrates that this enzyme can protect in vivo against the damage produced by the addition of H₂O₂.     

Lactic acid bacteria, their metabolic products and interference with microbial growth

Abstract: Lactic acid bacteria produce a variety of metabolic products that are capable of interfering with the growth of other microbes. These bacterial end products have been applied to food systems to prevent the growth of certain undesirable bacteria. The following review will discuss the successful application of several of the metabolic products produced by lactic acid bacteria in food systems.     

Bioprospecting for microbial endophytes and their natural products.

Endophytic microorganisms are to be found in virtually every plant on earth. These organisms reside in the living tissues of the host plant and do so in a variety of relationships, ranging from symbiotic to slightly pathogenic. Because of what appears to be their contribution to the host plant, the endophytes may produce a plethora of substances of potential use to modern medicine, agriculture, and industry. Novel antibiotics, antimycotics, immunosuppressants, and anticancer compounds are only a few examples of what has been found after the isolation, culture, purification, and characterization of some choice endophytes in the recent past. The potential prospects of finding new drugs that may be effective candidates for treating newly developing diseases in humans, plants, and animals are great.     

Quantitative analysis of pathways of methionine metabolism and their regulation in lemna

Individual rates of metabolism of the sulfur, methyl, and 4-carbon moieties of methionine were estimated in Lemna paucicostata Hegelm. 6746 growing under standard conditions, and used to quantitate pathways of methionine metabolism. Synthesis of S-adenosylmethionine (AdoMet) is the major pathway for methionine metabolism, with over 4 times as much methionine metabolized by this route as accumulates in protein. More than 90% of AdoMet is used for transmethylation. Methyl groups of choline, phosphatidylcholine, and phosphorylcholine are major end products of this pathway. Flux through methylthio recycling is about one-third the amount of methionine accumulating in protein. Spermidine synthesis accounts for at least 60% of the flux through methylthio recycling. The results obtained here, together with those reported for methionine-supplemented plants (Giovanelli, Mudd, Datko 1981 Biochem Biophys Res Commun 100: 831-839), indicate that methionine supplementation reduced methylneogenesis by no more than the small amount expected from the reduced entry of sulfate sulfur into methionine (Giovanelli, Mudd, Datko, 1985 Plant Physiol 77: 450-455). Methionine supplementation had no significant effect on transmethylation or methylthio recycling. The combined data provide the first comprehensive estimates of the quantitative relationships of major pathways for methionine metabolism and their control in plants.     

Mechanism of adhesion maintenance by methionine sulphoxide reductase in Streptococcus gordonii

Methionine sulphoxide reductase maintains adhesin function during oxidative stress. Using Streptococcus gordonii as a model, we now show the mechanistic basis of adhesin maintenance provided by MsrA. In biofilms, S. gordonii selectively expresses the msrA gene. When the wild-type strain was grown with exogenous hydrogen peroxide (H(2)O(2)), msrA-specific mRNA expression significantly increased, while acid production was unaffected. In the presence of H(2)O(2), a msrA-deletion mutant (ΔMsrA) showed a 6 h delay in lag phase growth, a 30% lower yield of H(2)O(2), significantly greater inhibition by H(2)O(2) on agar plates (reversed by complementation), 30% less adhesion to saliva-coated hydroxyapatite, 87% less biofilm formation and an altered electrophoretic pattern of SspAB protein adhesins. Using mass spectrometry, methionine residues in the Met-rich central region of SspB were shown to be oxidized by H(2)O(2) and reduced by MsrA. In intact wild-type cells, MsrA colocalized with a cell wall-staining dye, and MsrA was detected in both cell wall and cytosolic fractions. To maintain normal adhesion and biofilm function of S. gordonii in response to exogenous oxidants therefore msrA is upregulated, methionine oxidation of adhesins and perhaps other proteins is reversed, and adhesion and biofilm formation is maintained.     

The microbial biosynthesis of methionine

1. The enzymes leading to the methylation of homocysteine have been examined in three micro-organisms: a cobalamin-producing bacterium, Bacillus megaterium; a yeast, Candida utilis; and a basidiomycete fungus, Coprinus lagopus. The yeast and the fungus contain negligible endogenous cobalamin. 2. Extracts of each organism catalyse C(1)-transfer from serine to homocysteine with a polyglutamate folate coenzyme. 3. The enzymes generating the methyl group of methionine from C-3 of serine have similar properties in each case, but different mechanisms of homocysteine transmethylation from 5-methyltetrahydrofolates were found. 4. B. megaterium contains an enzyme with properties suggestive of a vitamin B(12)-dependent homocysteine transmethylase, whereas Cand. utilis and Cop. lagopus transfer the methyl group by a reaction characteristic of the cobalamin-independent mechanism established for Escherichia coli. 5. The specificity of each transmethylase for a 5-methyltetrahydropteroylpolyglutamate is consistent with the results of analyses of endogenous folates in these organisms, which showed only conjugated forms. 6. None of the extracts catalysed methionine production from S-adenosylmethionine and homocysteine. 7. These results are compared with results now available for methionine synthesis in other organisms, which show a considerable diversity of mechanisms.     

Chromatographic separation of multiple renin substrates in women: effect of pregnancy and oral contraceptives

Chromatographic separation of multiple renin substrates was carried out to study the effect of pregnancy and oral contraceptives (OCs) in women. Blood plasma proteins were separated on a Sephadex G-200 column and the amount of renin substrate in each fraction was determined. The molecular weight of the renin substrate peaks was estimated by comparison with the elution position of proteins of known molecular weight. Normal nonpregnant women revealed the presence of 2 renin substrates: a major 1 with a molecular weight of 65000 and 1 amounting to 3-5% of the quantity of the major peak with a molecular weight of 450000-500000. In women taking OCs the same 2 peaks were seen but the quantity of each was increased markedly. In pregnant women at term, a 3rd renin substrate of intermediate size was seen (molecular weight 350000 daltons). The quantities of the intermediate and low molecular weight renin substrates were found to be greatly increased in pregnancy.