Strategy towards the enantioselective synthesis of schiglautone A

ISSN:1477-0520ISSN:1477-053

Activation, structure, biosynthesis and bioactivity of glidobactin-like proteasome inhibitors from Photorhabdus laumondii

The glidobactin-like natural products (GLNPs) glidobactin A and cepafungin I have been reported to be potent proteasome inhibitors and are regarded as promising candidates for anticancer drug development. Their biosynthetic gene cluster (BGC) plu1881–1877 is present in entomopathogenic Photorhabdus laumondii but silent under standard laboratory conditions. Here we show the largest subset of GLNPs, which are produced and identified after activation of the silent BGC in the native host and following heterologous expression of the BGC in Escherichia coli. Their chemical diversity results from a relaxed substrate specificity and flexible product release in the assembly line of GLNPs. Crystal structure analysis of the yeast proteasome in complex with new GLNPs suggests that the degree of unsaturation and the length of the aliphatic tail are critical for their bioactivity. The results in this study provide the basis to engineer the BGC for the generation of new GLNPs and to optimize these natural products resulting in potential drugs for cancer therapy

A collaborative journey towards the late‐stage functionalization of added‐value chemicals using engineered halogenases

In 2017, two companies, Novartis AG and Syngenta AG, joined forces with the group of Prof. Buller, head of the Competence Center for Biocatalysis (CCBIO), to tackle the challenge of enzymatic late-stage halogenation. This biocatalytic method was considered to provide a more sustainable approach to late-stage halogenation of complex molecules than traditional synthetic approaches. Using machine-learning guided protein engineering, α-ketoglutarate dependent halogenases were evolved into versatile catalysts capable of selectively chlorinating inactivated C−H bonds. Structurally diverse molecules, namely an analogue of martinelline as well as two members of the soraphen natural product family, were enzymatically chlorinated at two distinct positions in a regio- and stereoselective manner, thus demonstrating the synthetic usefulness of such a strategy. As part of our three-year collaboration, flavin-dependent halogenases were also studied

Sequential Inactivation of Gliotoxin by the <i>S</i>‑Methyltransferase TmtA

The epipolythiodioxopiperazine (ETP) gliotoxin mediates toxicity via its reactive thiol groups and thereby contributes to virulence of the human pathogenic fungus <i>Aspergillus fumigatus</i>. Self-intoxication of the mold is prevented either by reversible oxidation of reduced gliotoxin or by irreversible conversion to bis­(methylthio)­gliotoxin. The latter is produced by the <i>S</i>-methyltransferase TmtA and attenuates ETP biosynthesis. Here, we report the crystal structure of TmtA in complex with <i>S</i>-(5′-adenosyl)-l-homocysteine. TmtA features one substrate and one cofactor binding pocket per protein, and thus, bis-thiomethylation of gliotoxin occurs sequentially. Molecular docking of substrates and products into the active site of TmtA reveals that gliotoxin forms specific interactions with the protein surroundings, and free energy calculations indicate that methylation of the C10a-SH group precedes alkylation of the C3-SH site. Altogether, TmtA is well suited to selectively convert gliotoxin and to control its biosynthesis, suggesting that homologous enzymes serve to regulate the production of their toxic natural sulfur compounds in a similar manner

Algorithm-aided engineering of aliphatic halogenase WelO5* for the asymmetric late-stage functionalization of soraphens

Late-stage functionalization of natural products offers an elegant route to create novel entities in a relevant biological target space. In this context, enzymes capable of halogenating sp3 carbons with high stereo- and regiocontrol under benign conditions have attracted particular attention. Enabled by a combination of smart library design and machine learning, we engineer the iron/α-ketoglutarate dependent halogenase WelO5* for the late-stage functionalization of the complex and chemically difficult to derivatize macrolides soraphen A and C, potent anti-fungal agents. While the wild type enzyme WelO5* does not accept the macrolide substrates, our engineering strategy leads to active halogenase variants and improves upon their apparent kcat and total turnover number by more than 90-fold and 300-fold, respectively. Notably, our machine-learning guided engineering approach is capable of predicting more active variants and allows us to switch the regio-selectivity of the halogenases facilitating the targeted analysis of the derivatized macrolides’ structure-function activity in biological assays

CCDC 901386: Experimental Crystal Structure Determination

Related Article: P.Mombelli,C.Le Chapelain,N.Munzinger,E.Joliat,B.Illiarov,W.B.Schweizer,A.K.Hirsch,M.Fischer,A.Bacher,F.Diederich|2013|Eur.J.Org.Chem.|2013|1068|doi:10.1002/ejoc.20120146

CCDC 901385: Experimental Crystal Structure Determination

Related Article: P.Mombelli,C.Le Chapelain,N.Munzinger,E.Joliat,B.Illiarov,W.B.Schweizer,A.K.Hirsch,M.Fischer,A.Bacher,F.Diederich|2013|Eur.J.Org.Chem.|2013|1068|doi:10.1002/ejoc.20120146