INVESTIGADORES
FRIES Alexander Erich
artículos
Título:
Alteration of the Route to Menaquinone towards Isochorismate-Derived Metabolites
Autor/es:
FRIES, ALEXANDER; MAZZAFERRO, LAURA S.; GRÜNING, BJÖRN; BISEL, PHILIPPE; STIBAL, KARIN; BUCHHOLZ, PATRICK C. F.; PLEISS, JÜRGEN; SPRENGER, GEORG A.; MÜLLER, MICHAEL
Revista:
CHEMBIOCHEM
Editorial:
WILEY-V C H VERLAG GMBH
Referencias:
Año: 2019 vol. 20 p. 1672 - 1672
ISSN:
1439-4227
Resumen:
Chorismate and isochorismate constitute branch-point intermediates in the biosynthesis of many aromatic metabolites in microorganisms and plants. To obtain unnatural compounds, we modified the route to menaquinone in Escherichia coli. We propose a model for the binding of isochorismate to the active site of MenD ((1R,2S, 5S,6S)-2-succinyl-5-enolpyruvyl-6-hydroxycyclohex-3-ene-1-carboxylate (SEPHCHC) synthase) that explains the outcome of the native reaction with α-ketoglutarate. We have rationally designed variants of MenD for the conversion of several isochorismate analogues. The double-variant Asn117Arg?Leu478Thr preferentially converts (5S,6S)-5,6-dihydroxycyclohexa-1,3-diene-1-carboxylate (2,3-trans-CHD), the hydrolysis product of isochorismate, with a >70-fold higher ratio than that for the wild type. The single-variant Arg107Ile uses (5S,6S)-6-amino-5-hydroxycyclohexa-1,3-diene-1-carboxylate (2,3-trans-CHA) as substrate with >6-fold conversion compared to wild-type MenD. The novel compounds have been made accessible in vivo (up to 5.3 g L−1). Unexpectedly, as the identified residues such as Arg107 are highly conserved (>94 %), some of the designed variations can be found in wild-type SEPHCHC synthases from other bacteria (Arg107Lys, 0.3 %). This raises the question for the possible natural occurrence of as yet unexplored branches of the shikimate pathway.