Methionine and derivatives: Exploring chirality at sulfur

Abstract

Chiral molecules, both small and large, play important roles in biochemistry, molecular biology, and related biosciences. The central position of carbon atoms in determining chirality has received considerable attention. Less well known is the fact that chirality is also possible for sulfur atoms in certain compounds. This article introduces the role of sulfur chirality in metabolic reactions by considering methionine derivatives with chiral sulfur atoms, S-adenosylmethionine and its decarboxylated product, methionine S-oxide, and methionine sulfoximine.

Of the 22 amino acids incorporated into protein by the ribosomal process, methionine must be the superstar. It plays a fundamental role in protein biosynthesis and structure and has countless functions in general metabolism. Met derivatives also participate in cell signaling, as for instance the role of formyl-Met-Leu-Phe-OH in responses of neutrophils. Moreover, Met is the precursor for the biosynthesis of dimethyl sulfide and methanethiol, major components of the group known as volatile organic sulfur compounds [1]. These materials are involved in global warming, production of acid rain, and cloud nucleation. Some 40 million metric tonnes of dimethyl sulfide are released to the atmosphere annually. Perhaps it can be claimed that Met influences our weather as well as our physiology.

Together with some other amino acids, Met is characterized by the -CH₃-S- group; however, Met is the only such material found in protein. It is a sulfide, R¹-S-R², where R¹ = CH₃ and R² = (CH₂)₂CH(NH₂)COOH. Sulfides are reactive compounds readily converted to sulfoxides, R¹-SO-R², and sulfones, R¹-SO₂-R², by oxidation, to sulfonium salts, R¹R²R³S⁺, by addition of a cation, e.g. CH₃⁺, and to sulfoximines, R¹-SO(NH)-R², by complex processes. A fact that usually receives little attention in the teaching of biochemistry/molecular biology is that some compounds derived from Met contain a chiral sulfur atom: they are S-adenosylmethionine and related materials (sulfonium salts), Met-S-oxide (a sulfoxide) and Met sulfoximine. Chiral sulfur compounds have long been of interest to organic chemists [2], being first described more than a century ago. As with chiral carbon compounds, enantioselective and diastereoselective interactions can occur with other chiral substances such as enzymes and receptors, thus influencing physiological events.

ADENOSYLMETHIONINE AND RELATED MATERIALS

S-Adenosylmethionine (AdoMet),11 a very versatile metabolite, is the second most widely used enzyme substrate after ATP [3]. Famous as the almost universal donor of methyl groups to both small and macro molecules, it also functions as an amino donor (transamination of 7-amino-8-ketopelargonic acid), as a propylamine donor (synthesis of spermidine and spermine), and in the formation of ethylene (in plants). It is a typical sulfonium salt, R¹R²R³S⁺, where R¹ = CH₃, R² = adenosine, and R³ = (CH₂)₂CH(NH₂)COOH. Structurally, such compounds form a pyramidal arrangement of the three R groups, and there is a lone electron pair also associated with the sulfur atom. Whereas a pyramidal inversion is possible, many compounds have an inversion barrier of about 25–30 kcal mol⁻¹ so that if R¹ ≠ R² ≠ R³, stable enantiomers may be formed for sulfonium salts; the compounds show chirality dependent on sulfur. In addition to a chiral sulfur at the sulfonium pole, AdoMet has one chiral carbon at the α amino position and four further chiral carbons in the ribose moiety of adenosine. Hence there is a total of 2⁶ stereoisomeric possibilities. If the ribose carbons are considered invariant, there are four stereoisomeric forms deriving from the α amino carbon and the sulfur atom. If the former has the usual L(S) configuration, there are two diastereoisomers to be considered.

That they could behave differently in enzymatic reactions was established as follows [4]. AdoMet obtained by action of rabbit liver L-methionine adenosyltransferase, EC 2.5.1.6, was 100% utilized as a methyl donor for creatine formation by guanidinoacetate N-methyltransferase, EC 2.1.1.2. However, AdoMet obtained by chemical methylation of S-adenosylhomocysteine was only used to the extent of 50% under the same experimental conditions. In other words, the chemically synthesized material was a mixture of two isomers, one of which did not function as a methyl transfer material.

Configurations at pyramidal centers can be specified by the usual Cahn-Ingold-Prelog procedure; the “absent” fourth ligand is the electron lone pair, assigned an atomic number of zero for the purpose of determining the priority sequence. Thus, for the AdoMet sulfur, the priority sequence is as follows: (The superscript numbers refer to carbon atoms of the ribose moiety.) In compounds such as AdoMet with both chiral sulfur and carbon compounds, the configurational descriptors, (R) and (S), are modified by use of subscript s for sulfur and c for carbon. Care is always necessary not to confuse S indicating substitution at sulfur with the configurational descriptor. Assuming invariance at the ribose chiral carbons, enzymatically synthesized AdoMet is (S_cS_s)-S-adenosylmethionine (R = COOH; Fig. 1A). The determination of the absolute configuration at sulfur for enzymatically active AdoMet by a group headed by Sir John W. Cornforth (Nobel Laureate in Chemistry, 1975) was an outstanding achievement [5]. With the possible exception of a poorly documented example, methyl transferase enzymes are specific for this (S_cS_s) diastereoisomer. The (S_cR_s) isomer is an inhibitor of methylases. Methods have been developed for the separation and assay of the diastereoisomers [6].

For the biosynthesis of polyamines such as spermidine and spermine, decarboxylation of AdoMet by S-adenosylmethionine decarboxylase, EC 4.1.1.50, is the first step. It is stated [7] that the decarboxylated metabolite, dcAdoMet, is formed with retention of configuration at the sulfur and thus has the (S_s) structure (R = H; Fig. 1A). Spermidine synthase is specific for this isomer.

1-Aminocyclopropane-1-carboxylate, required as an intermediate in ethylene biosynthesis, is formed from AdoMet by action of 1-aminocyclopropane-1-carboxylate synthase, EC 4.4.1.14. The enzyme from tomato uses only (S_cS_s)-AdoMet as substrate, and the (S_cR_s) isomer is a strong inhibitor [8, 9].

METHIONINE S-OXIDE

It is recommended that the mono-oxidation product of Met be named as methionine S-oxide (methionine oxide), because the term methionine sulfoxide might imply the presence of a further sulfur atom in addition to that already present in Met [10]. This recommendation has received less application than it deserves, but the recommended symbol, MetO, is one of several in use. Because MetO contains a chiral carbon and a chiral sulfur, four stereoisomers are possible. MetO (Fig. 1B) isolated from the blowfly, Phormia regina, has the (S_cS_s) configuration [11, 12]. Free and peptide-bound Met is readily oxidized, and the process in proteins is often accompanied by structural and functional changes [13]. The process can be reversed by MetO reductases, which thus function as in vivo antioxidants. There is a vast literature on this subject, and moreover, the terminology is unusually confusing. Only a brief account is possible here emphasizing structural considerations.

Two reductase types are distinguished: methionine-S-oxide reductase, EC 1.8.4.5, does not react with protein-bound MetO, whereas protein-methionine-S-oxide reductase, EC 1.8.4.6, reacts with both free and bound MetO. It is generally accepted that MetO present in proteins is a mix of (S_cS_s) and (S_cR_s) isomers. One much-studied reductase, often termed MsrA, is specific for reduction of (S_cS_s)-MetO in both free and protein-bound forms [14, 15]. When the msrA gene was overexpressed in Drosophila, the lifetime of the flies was extended, and they were more active physically with normal food intake and body weight [16]. The authors expressed interest in determining whether “over-expression of MSRA extends lifespan in mammals including humans,” an intriguing possibility involving a chiral sulfur compound!

A second enzyme type, often termed MsrB and specific for (S_cR_s)-MetO, is present in several organisms including bacteria, yeast, fruit fly, and mammals [17]. One interesting mammalian protein with MsrB activity is selenoprotein R; this enzyme contains the amino acid selenocysteine [18].

METHIONINE SULFOXIMINE

Methionine sulfoximine was originally isolated from flour treated with nitrogen trichloride (“agene”) to improve baking properties. Because it was toxic to dogs and ferrets, this process was discontinued. It has now been isolated from fresh seeds of Cnestis palala, a woody plant of the tropics [19]. The biologically active form (Fig. 1C) has the (S_cS_s) configuration [20]. The structural arrangement producing chirality in a sulfoximine is different from that involved for sulfonium salts and sulfoxides. The molecular arrangement of four groups around the sulfur is tetrahedral. Note that sulfones, R¹–SO₂–R², R¹ ≠ R², also are tetrahedral but chirality can only be observed by using two oxygen isotopes as in R¹–S¹⁶O¹⁸O–R², R¹ ≠ R².

Methionine sulfoximine has been extensively employed as an inhibitor of glutamine synthetase and γ-glutamylcysteine synthetase [21]. Glutamine synthetase in the presence of ATP adds a phosphate group at the nitrogen atom. Only the (S_cS_s) isomer undergoes this phosphorylation, but both (S_cS_s) and (S_cR_s) forms bind reversibly to a single subunit site [22]. Analogs in which the CH₃ group is replaced by other alkyl units (ethyl, propyl, butyl) have variable actions on these two enzymes. The butyl analog, S-(n-butyl)homocysteine sulfoximine, buthionine sulfoximine, is the only one for which stereochemical information is available [23]. The (S_cS_s) isomer was bound to and inhibited γ-glutamylcysteine synthetase as the phosphate, but buthionine sulfoximine did not inhibit glutamine synthetase.

FINAL COMMENTS

Whereas this article has focused on Met and its derivatives, other compounds with chiral sulfur atoms have interesting biological properties. Many cysteine derivatives in the genus Allium are sulfoxides, and sulfoxide structures are also of concern for biotin, lipoic acid, penicillins and cephalosporins, amatoxins, and some other natural products (e.g. leinamycin). If the topic seems a little academic to students (Oh, no! Not stereochemistry again!), one practical, economic consequence of sulfur chirality might grab their attention. The proton pump-inhibiting drug, Prilosec (omeprazole), now available over the counter in the United States, has had great success. Structurally, it is a sulfoxide with a single chiral sulfur center, and it is marketed as a racemate. The single (S) enantiomer form, Nexium (esomeprazole), has improved clinical properties and has also been a great financial success [24]. Because there are other sulfoxide drugs, perhaps another “racemic switch” is inevitable.

Figure Fig. 1.. Chiral sulfur compounds. A, R = COOH, the enzymatically active isomer of AdoMet. The lone electron pair (data not shown) is behind the plane of the paper. Hence, the configurational descriptor at sulfur can be read directly as (S) by observing that the sequence, ribose unit to Met unit to CH₃, is left-handed. Also, R = H, the decarboxylation product of AdoMet. In the decarboxylation, chirality is lost at the original α carbon of the methionine unit. B, methionine S-oxide, shown as the (S_cS_s) isomer. The lone pair electrons on the sulfur atom are indicated by e. C, methionine sulfoximine shown as the (S_cS_s) isomer.

Acknowledgements

E. J. Behrman (Ohio State University) is thanked for helpful suggestions. Readers might like to know that recent research on MetO will be extensively discussed in a forthcoming special issue of Biochimica et Biophysica Acta: Proteins and Proteomics. Some articles are already available on-line as of December 2004. Thanks to C. Glaser and H. Weissbach for this information.

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  The abbreviation used is: AdoMet, S-adenosylmethionine.