Methionine sulfonium salts

This enzyme [EC 3.3.1.2], also referred to as -adenosyl-methionine cleaving enzyme and methylmethionine-sulfonium-salt hydrolase, catalyzes the hydrolysis of -adenosylmethionine to produce methylthioadenosine and homoserine. The enzyme will also convert methyl-methionine sulfonium salt to dimethyl sulfide and homoserine. 

The involvement of methionine sulfonium salts in these chemical cleavage reactions may stimulate experiments to show the significance of special methionine residues in the active centers and the catalytic sites of enzymes. Such an important role for a particular methionine has been postulated for the enzymes phosphoglucomutase and chymotrypsin on the basis of photooxidation studies (Ray et al., 1960). 

Wong, F.F. and Carson, J.F., 1966. Isolation of S-methyl methionine sulfonium salt from fresh tomatoes. J. Agric. Food Chem., 14 247—249. 

Analysis of samples showed that concentrations of methanethiol and dimethyl disulfide were lower in caraway- and TBHQ-treated broccoli florets than in control samples (Table V). Dimethyl tiisulfide was not detected in the refrigerated samples which indicated that its contribution to the objectionable odors of modified atmosphere stored broccoli florets was minimal. Dimethyl sulfide which has a precursor (Sr methyl methionine sulfonium salt 34) different from that for methanethiol-related compounds was present in all samples. 

A naturally occurring sulfonium salt S adenosylmethionme (SAM) is a key sub stance in certain biological processes It is formed by a nucleophilic substitution m which the sulfur atom of methionine attacks the primary carbon of adenosine triphosphate dis placing the triphosphate leaving group as shown m Figure 16 7... 

Divalent sulfur compounds are achiral, but trivalent sulfur compounds called sulfonium stilts (R3S+) can be chiral. Like phosphines, sulfonium salts undergo relatively slow inversion, so chiral sulfonium salts are configurationally stable and can be isolated. The best known example is the coenzyme 5-adenosylmethionine, the so-called biological methyl donor, which is involved in many metabolic pathways as a source of CH3 groups. (The S" in the name S-adenosylmethionine stands for sulfur and means that the adeno-syl group is attached to the sulfur atom of methionine.) The molecule has S stereochemistry at sulfur ana is configurationally stable for several days at room temperature. Jts R enantiomer is also known but has no biological activity. 

The sulfur atom of methionine residues may be modified by formation of sulfonium salts or by oxidation to sulfoxides or the sulfone. The cyanosulfonium salt is not particularly useful for chemical modification studies because of the tendency for cyclization and chain cleavage (129). This fact, of course, makes it very useful in sequence work. Normally, the methionine residues of RNase can only be modified after denaturation of the protein, i.e., in acid pH, urea, detergents, etc. On treatment with iodoacetate or hydrogen peroxide, derivatives with more than one sulfonium or sulfoxide group did not form active enzymes on removal of the denaturing agent (130) [see, however, Jori et al. (131)]. There was an indication of some active monosubstituted derivatives (130, 132). 

With methyl iodide as the alkylating reagent, Link and Stark (133) prepared a monosubstituted sulfonium salt. Methionine 29 was strongly indicated as the principal site of modification. The activity of the derivative toward C>p was the same as RNase-A, and the methylation reaction was not affected by competitive inhibitors such as 2 (3 )-UMP. 

Methionine 13 in S-peptide has been modified by oxidation to the sul-fone or by conversion to a sulfonium salt with either iodoacetic acid or iodoacetamide (138). There is a dramatic lowering of the peptide-protein binding constant for all of the derivatives, but the complexes when formed appear to have nearly normal catalytic activity. The X-ray structure does not appear to permit the normal sulfur location with the sulfonium salts. Sterically, Met 13 can be moved by rotation about the carbon a-carbon / bond so that the residue sticks out into the solvent. This can be done without any major change in the conformation of the rest of the peptide. Thus the active site could be maintained undisturbed while the contribution of Met 13 to the S-peptide S-protein association would be lacking. 

The first metal(salen) complex investigated as an asymmetric phase-transfer catalyst was methionine-derived sulfonium salt 29. It was anticipated that the sulfonium salt... 

Sulfonium salts are common alkylating agents in biological systems. For example, ATP activation of methionine forms the sulfonium salt S-adenosyl methionine (SAM), a biological methylating agent. 

In view of these many precedents it is surprising that the intramolecular displacement of the sulfur function of methionine derivatives has not been utilized for selective cleavage of methionine peptides. The method has only recently been applied to a number of methionine peptides which according to the following scheme (CXX-CXXIII) were first converted to sulfonium salts and then subjected to intramolecular (imino) lactonization and hydrolysis by short heating in water (Lawson et al., 1961). Table XV shows the strong influence of the nature of the added alkyl group on the yield in... 

Aminocyclopropane carboxylic acid (6) has been detected in several plant tissues a procedure for preparing 6 from agricultural wastes, by extraction with a diluted solution of sulfosalicylic acid, has been described . 6 was established to be an intermediate product in ethylene biosynthesis " . Ethylene acts as a phytohormone which is involved in many metabolic processes in plants, e.g. in ripening, in stress situations or after wounding (see review and references cited therein). Natural 6 is formed from methionine via sulfonium salt (640) only S,S-(640) acted as a substrate for aminocyclopropanecarbo-xylate synthase, the S,R and R,R isomers of 640 were inactive as substrates . 6 can be... 

The method described above gives direct analysis of methionine sulfoxide content in proteins. Another method makes use of carboxy-methylation of methionine at acid pH to give the carboxymethyl-sulfonium derivative ( 3.5). Methionine sulfoxide, which is not affected by the carboxymethylation reaction, is then oxidized to the sulfone which is stable to acid hydrolysis and can easily be quantitated. This is possible because methionine carboxymethyl-sulfonium salts are not affected by performic acid oxidation, although they are degraded by acid hydrolysis. Therefore, the methionine sulfone content is equal to the methionine sulfoxide content plus any sulfone that may have been initially present (shown by analysis before oxidation). 

Naider and Bohak (1972) have found that incubation of sulfonium salts with thiols leads to the regeneration of methionine. Complete recovery of methionine was obtained upon incubation of the S-carboxyamidomethylmethionyl sulfonium salt with 0.12 M mercapto-ethanol at pH 8.9 for 24 hr at. 30°C. In contrast, the S-carboxymethyl-methionyl sulfonium salt is regenerated to only a minor extent under these conditions. 

The derivatives of methionine formed by haloketones are not stable to the usual conditions of acid hydrolysis. These sulfonium salts are degraded in three different ways. Some methionine is regenerated, some homoserine and homoserine lactone is formed and possibly the homocysteine derivative of the general structure indicated below is produced where X represents the rest of the affinity label. 

The degradation product of the sulfonimn salt formed in the greatest yield after acid hydrolysis is undoubtedly a function of the structure of the affinity label. Generally, if the total amino acid composition of the modified protein was determined, low yields of methionine and increased yield of homoserine would be indicative, but certainly not proof of, the modification of a methionine residue. Sulfonium salts of methionine are generally not oxidized by hydrogen peroxide (Sigman and Blout 1967). 

Only four naturally occurring sulfonium salts are known. Two examples are dimethyl-p-propiothetin (8) and S-methyl-L-methionine (9) (Figure 2). Propiothetin (8) occurs in algae, plankton, fish and molluscs, and S-methyl-L-methionine (9) in cabbage, celery and other vegetables. 

Hi) Methionine Sulfoxide and Sulfone. Under controlled conditions, mild oxidizing agents convert methionine to its sulfoxide. It can also be formed by simple exposure of peptides and proteins to air for a long period of time. This derivative is unstable to acid and only partial recovery is observed after such treatment, but accurate estimates can be made from alkaline or enzymatic hydrolyses (Means and Feeney, 1971). It can also be determined by difference, a method which takes advantage of the fact that the sulfoxide does not react with alkylating agents and that sulfonium salts cannot be further oxidized to the sulfone (Neumann et a/., 1962). 

The reaction mixture (final volume 0.34 ml) contained (in /umol and including compounds added with the enzyme) potassium phosphate, pH 6.3, 102 Tris, pH 7.6, 5 sucrose, 13 GSH, 16 2-mercaptoethanol, 0.7 Na EDTA, 0.5. Methyl donors were added at the specified final concentrations. The various stereoisomeric forms of S-adenosylmethionine were prepared as described by de la Haba el al. (1959) using adenosylmethionine cyclotransferase (E.C. 2.5.1.4) to resolve (/(S)-S-adenosyl-L-methionine to (f )-5-adenosyl-L-methionine. The reaction was started by addition of enzyme dissolved in Tris buffer, pH 7.6, 25 mM, containing Na EDTA, 1 mM and 2-mercaptoethanol, 7 mM, and partially purified from an extract of cabbage leaves by successive ammonium sulfate precipitation (45-60% saturation), gel filtration (Sephadex G-25) and DEAE-cellulose column chromatography. After incubation at 26°C for 1 h, each reaction mixture was diluted with 0.76 ml cold water, and a 1.0 ml aliquot was immediately added to a column of Dowex-50(NHJ) 1 x 3 cm. Unreacted [ C]methionine was removed by washing the column with 25 ml water. The 5-[" C]methylmethionine sulfonium salt was then eluted with 8 ml 0.6 N NH OH. Radioactivity was determined by scintillation spectrophotometry (Mudd et al., 1965). A blank of 540 cpm has been subtracted from each value. The low activity with (R)-5-adenosyl-L-methionine may have been due to the presence of a small residual amount of (5)-5-adenosyl-L-methionine (de la Haba eta/., 1959). 

It should be noted that the sulfur atom in methionine sulfoxide is a new chiral center, hence a pair of diastereosimers is present in the intermediates. This is a minor complication that is absent from an alternative method for the masking of the thioether its more recently proposed conversion to a tert.sulfonium salt by the reaction of an activated N-acyl derivative of the amino acid with methyl p-toluenesulfonate ... 

Electrophilic substitution of aromatic nuclei in tyrosine and tryptophan side chains has frequently been reported in connection with acidolytic removal of blocking groups. C-Benzylation and tert.butylation of the tyrosine side chain and N-alkylation of the indole nucleus in tryptophan are often attributed to the alkyl cations generated in the reaction. This common side reaction is caused, however, mainly by the alkylating agents formed in the process, such as benzyl bromide or tert.butyl trifluoroacetate. The same is true for the S-alkylation of the methionine side chain. Conversion of the thioether to a sulfonium salt can... 

This side reaction is reversible. As already mentioned in connection with the protection of the methionine side chain (Chapter VI) the thioether can be regenerated from the ternary sulfonium salts by treatment with mercaptanes under mildly basic conditions. Nevertheless, this long overlooked alkylation should serve as warning against the indiscriminate use of scavangers. The more acid stable thioanisole or the equally efficient 4-methylthiophenol are less likely to cause similar problems. 

It is well known that sulfonium salts derived from methionine may decompose in many ways. Electron-withdrawing groups on the sulfur atom promote the breakdown of the sulfonium salt by bimolecular or intramolecular nucleophilic displacement. In the case of acetyl-methionine sulfonium derivatives (21) the displacement reaction takes place with the participation of the C-carbonyl group via a 1,5-interactipn, resulting in elimination of the dialkyl sulfide and formation of homoserine iminolactone (22) which on hydrolysis gives homoserine lactone (23) (reaction 17) . Best results are obtained when the reac-... 

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