research highlights

Biocatalysis EDDS lyase sweetening your day Angew. Chem. Int. Ed. http://doi.org/dc7g (2019)

Oneofthe most used artificial low-calorie sweeteners Derivativesof aspartame areupto 100-foldsweeter

O O O O O O O O O O H H H2N N N N N N H H H HO HO HO

O O O Aspartame Neotame Advantame 200 timessweeter thansucrose 7,000–13,000 times sweeter than sucrose 20,000times sweeter than sucrose

O O O O H O O R N O OH H H N OH EDDSlyase R=CH C(CH ) : 2 R N + R NH2 HO 2 3 3 N HO N-(3,3-dimethylbutyl)-L-aspartic: Peptidase H O HO O R=CH2CH2[3-OH)-4-(OCH3)C6H3]: N-[3-(3-hydroxy-4-methoxyphenyl)propyl]-L-aspartic O

Artificial low-calorie sweeteners are widely (pictured, bottom). First, the authors showed Favourable apolar–apolar contacts of the used in foodstuffs with the aim of reducing that wild-type EDDS lyase allows for the mutated residues with the 3,3-dimethylbutyl sugar consumption. One of the most synthesis of N-(3,3-dimethylbutyl)-l-aspartic moiety lead to a stronger binding and used artificial sweeteners is the dipeptide acid and N-[3-(3-hydroxy-4-methoxyphenyl) presumably a more productive binding of the aspartame. Neotame and advantame are propyl]-l-aspartic acid under excess of the amine compound. recently approved sugar substitutes that are amine substrate with high conversion and Lyases are enzymes that usually catalyse obtained via derivatization of aspartame excellent enantioselectivity, but reaction the breaking of bonds. This work is another with branched N-alkyl- or N-arylalkyl times were long (7 days). Therefore, to example that shows that engineering groups, respectively (pictured, top). These enhance the hydroamination activity of the reverse reaction of enzymes can be a modifications increase the sweetness — with EDDS lyase, protein engineering — based on powerful approach. Here, it allowed fast advantame being around 100 times sweeter the structure of EDDS lyase in complex with and easy access to optically pure precursors than aspartame. its natural substrate (S,S)-ethylenediamine- of valuable molecules that could find A biocatalytic asymmetric strategy for N,N′-disuccinic acid ((S,S)-EDDS) — was application in daily life. For example, the sustainable and step-economic synthesis applied. Two residues (Asp290 and Tyr320) with 0.05 mol% loading of the engineered of these compounds was lacking. Now, that were assumed to position the amine biocatalyst, the neotame precursor Poelarends and colleagues report that EDDS substrate in the active site were chosen N-(3,3-dimethylbutyl)-l-aspartic acid was lyase can convert fumaric acid to optically for mutation. The mutant EDDS(D290M/ produced with 96% conversion and >99% pure N-(3,3-dimethylbutyl)-l-aspartic acid Y320M) showed a 1,140-fold increase in enantiomeric excess after only 2.5 hours. and N-[3-(3-hydroxy-4-methoxyphenyl) activity over the wild-type enzyme. To propyl]-l-aspartic acid, which can potentially rationalize the effect of the mutations, crystal Jan-Stefan Völler be subsequently converted to neotame and structures of this enzyme variant were solved advantame, respectively, by a peptidase- and the neotame precursor was docked into Published online: 14 November 2019 catalysed amide-bond-coupling reaction the wild-type and mutant crystal structure. https://doi.org/10.1038/s41929-019-0389-1

948 Nature Catalysis | VOL 2 | NOVEMBER 2019 | 948 | www.nature.com/natcatal