Design of α-Keto Carboxylic Acid Dimers by Domain Recombination of Nonribosomal Peptide Synthetase (NRPS)-Like Enzymes

Abstract

Nonribosomal peptide synthetase (NRPS)-like enzymes comprising A–T–TE architectures catalyze the dimerization of α-keto carboxylic acids leading to the formation of hydroxybenzoquinones or lactones. Domain change experiments with five enzymes revealed that A and A–T domains of phenyl or 4-hydroxyphenyl pyruvate-using enzymes can be effectively used by the TE domains of other enzymes. Even the A and A–T domains of an indolyl hydroxybenzoquinone synthase were successfully recombined with TE domains of a phenyl and a 4-hydroxyphenyl pyruvate-activating enzyme.

Nonribosomal Peptides Produced by Minimal and Engineered Synthetases with Terminal Reductase Domains

“…However, a version of NRPS‐4 maintaining the T−R didomain from XtvB (NRPS‐5) results in an approximately 6.5‐fold lower production of both derivatives. Preservation of the natural A−T didomain has also been reported previously in engineered NRPS systems with A−T−TE architecture . Furthermore, the fusion point C‐terminal to the last helix of the T domain used in this work (Figure S23) was shown in our development of the XU concept to be applicable for introducing terminal C domains for peptide release .…”

Expanding the chemical diversity of an endophytic fungusBulgaria inquinans, an ascomycete associated with mistletoe, through an OSMAC approach

“…Chemical investigation of B. inquinans grown on solid Czapek medium resulted in the isolation of ve new natural products, namely butyrolactones (1-3, 5) and a new metabolite bearing a diol moiety (4) together with known compounds, such as the azaphilone pigment bulgarialactone B (14) 29 as well as phenylbutyrolactone IIa (15) 33,34 and xenofuranone B (17) 35 (Fig. 1).…”

“…The structures of the remaining known compounds isolated from B. inquinans were established as phenylbutyrolactone IIa (15), 33,34,59 xenofuranone B (17) 35,67 and bulgarialactone D (18), 30 based on their spectroscopic data as well as on a comparison with the literature.…”

Characterisation of ascocorynin biosynthesis in the purple jellydisc fungus Ascocoryne sarcoides

“…The metabolites produced by these enzymes contribute to a range of different biological activities such as phytotoxicity [ 1 ], quorum sensing [ 2 ] or the formation of protective pigments [ 3 ] to mention only a few. The domain structure of non-reducing NRPS-like enzymes consists of an adenylation (A), thiolation (T) and thioesterase (TE) domain [ 4 ] but they lack the condensation (C) domain that is vital for peptide bond formation in true non-ribosomal peptide synthetases. Thus, non-reducing NRPS-like enzymes are unable to produce peptide bonds and do not seem to use amino acids as substrates.…”

“…Thus, non-reducing NRPS-like enzymes are unable to produce peptide bonds and do not seem to use amino acids as substrates. Instead, in all NRPS-like enzymes characterised so far the A domain activates two identical aromatic alpha-keto acids that derive from either phenylalanine, tyrosine or tryptophan [ 4 ]. The T domain accepts the activated alpha-keto acids on its phosphopantetheine-loaded acyl carrier protein (ACP) and transfers the substrates to the TE domain that eventually performs a condensation reaction of the two substrate molecules [ 5 ].…”