Synthesis of 3‐arsonoalanine and its action on aspartate aminotransferase and aspartate ammonia‐lyase

Abstract

) -EJB 93 0201/3 D~-3-Arsonoalanine has been synthesized by the Strecker synthesis from the unstable compound arsonoacetaldehyde. It inactivates pig heart cytosolic aspartate aminotransferase and inhibits aspartate ammonia-lyase by competing with aspartate (KJK,,, 0.23). The fumarate analogue (0-3-arsonoacrylic acid and the malate analogue (RS)-3-arsonolactate also inhibit fumarate hydratase, competing with fumarate (K,/K, 1.8) and malate (Ki/Km 1.6) respectively. Attempted non-enzymic transamination of 3-arsonoalanine gave elimination of arsenite, in contrast with the transamination of 3-phosphonoalanine, which is either successful or leads to loss of phosphate.Phosphonoalanine (2-amino-3-phosphonopropionic acid) occurs naturally (Horiguchi, 1984) and is biologically transaminated (Roberts et al., 1968). Its transamination product, phosphonopyruvate, is the first product in the biosynthesis of the carbon-phosphorus bond and is formed by isomerization of phosphoenolpyruvate (Seidel et al., 1988; Bowman et al., 1988). The present study describes the synthesis of the arsenical analogue of phosphoalanine. Since a 2-oxoalkylarsonic acid is unstable (Lacoste et al., 1992), any transamination of 3-arsonoalanine to arsonopyruvate might be followed by release of arsenate ; thus it might poison organisms that take it up. Kamal (1989) explored many synthetic routes that might lead to arsonoalanine, but it was only after Lacoste et al. (1992) showed that arsonoacetaldehyde, although unstable, could be made in solution by the action of periodate on 2,3-dihydroxypropylarsonic acid, that we could make arsonoalanine from this aldehyde by a Strecker synthesis. We therefore report its synthesis and some of its properties, as well as its elimination of arsenite on attempted transamination to arsonopyruvate.Since arsonoalanine is an analogue of aspartate, we have tested it on two enzymes that use aspartate as substrate: aspartate aminotransferase, which was slowly inactivated, and aspartate ammonia-lyase, which was inhibited competitively. As aspartate ammonia-lyase and fumarate hydratase (fumarase) catalyse analogous reactions involving reversible trans-elimination of water or ammonium from (S)-malate or L-aspartate, respectively, with the formation of fumarate (Scheme l), we also studied the action of the fumarate analogue (E)-3-arsonoacrylate and the malate analogue 3-arsonolactate (3-arsono-2-hydroxypropionic acid) on fumarate hydratase. They also proved to be competitive inhibitors.

Arsenite release on enzymic transformation of arsonomethyl substrate analogues: a potentially lethal synthesis by glycerol-3-phosphate dehydrogenase

“…Repeated failures to isolate 4-hydroxy-3-oxobutylarsonic acid, the presumed product of the enzyme-catalysed oxidation of 3,4dihydroxybutylarsonic acid, followed by repeated failures to prepare it by methods similar to those used for the corresponding phosphonate [18], led us to try another method of preparation, starting with the transamination of the glutamate analogue 2amino-4-arsonobutyrate. Electrophoretic analysis of the reaction mixture showed that arsenite was eliminated, as it had been when Ali and Dixon [19] tried to transaminate the aspartate analogue 2-amino-3-arsonopropionate. Hence all the failures could be explained if 3-oxoalkylarsonic acids eliminated arsenite in aqueous solution.…”

“…Although the carbon-arsenic bond in alkylarsonic acids is formed from arsenite in the Meyer [8] reaction, breakage of this bond may occur at lower pH when arsenite becomes a better leaving group. The release of arsenite from benzylarsonic acid in HCl is known [20,21], as well as its release (see above) on attempted transamination of arsonoalanine [19].…”

“…The elimination of arsenite from 3-oxoalkylarsonic acids, but not of phosphite from 3-oxoalkylphosphonic acids, may be explained by the high stability of arsenite compared with arsenate, in contrast with the low stability of phosphite compared with phosphate. 2-Oxoalkylarsonic acids, exemplified by arsonoacetaldehyde [7,19] and 3-arsonopyruvate [23], are only moderately stable, but they release arsenate by hydrolysis, rather than arsenite by elimination. 2-Oxoalkylphosphonic acids can be hydrolysed in this way [24], as in the enzyme-catalysed hydrolysis of phosphonoacetaldehyde [25][26][27], but with much more difficulty.…”

Synthesis of 3-arsonopyruvate and its interaction with phosphoenolpyruvate mutase

“…Kamal [20] attempted the oxidation of tripropyl arsonolactate, using dicyclohexylcarbodi-imide with anhydrous orthophosphoric acid in dimethyl sulphoxide [21], but only arsenate was observed. Ali and Dixon [22] attempted the transamination of arsonoalanine using identical conditions to those employed by Sparkes et al [23] in the transmination of 3-phosphonoalanine to phosphonopyruvate with glyoxylate as amino group acceptor and Cu21 and pyridine as catalysts, but arsenite was released. Finally, Ali [24] treated bromopyruvic acid with alkaline arsenite, but arsonopyruvic acid was not formed.…”

“…They failed to isolate a solid sample because of its partial breakdown to arsenate on handling. Nevertheless, the solution could be used to synthesize arsonoalanine by a Strecker synthesis [22,24].…”