Flexible enzymatic activation of artificial polyketide extender units by Streptomyces cinnamonensis into the monensin biosynthetic pathway

Abstract

Polyketide natural products are of enormous relevance in medicine. The hit-rate in finding active compounds for the potential treatment of various diseases among this substance family of microbial origin is high. However, most polyketides require derivatization to render them suitable for the application. Of relevance in this field is the incorporation of artificial substances into the biogenesis of polyketides, hampered by both the microbial metabolism and the complexity of the enzymes involved. This manuscript describes the straightforward and selective biosynthetic incorporation of synthetic substances into a reduced polyketide and showcases a promising new enzyme to aid this purpose.

Exploring the Promiscuous Enzymatic Activation of Unnatural Polyketide Extender Units in Vitro and in Vivo for Monensin Biosynthesis

“…In order to explore the in vivo applicability of MatB_Sc, five malonate derivatives [2‐allyl, 2‐propyl, 2‐propargyl, 2‐butyl and 2‐(3‐chloropropyl)] were supplemented to a fermentation culture of S. cinnamonensis ATCC15413, assuming a co‐expression of MatB_Sc and the monensin PKS, as observed in the case of analogous premonensin fermentations . Five replicates were analyzed independently to allow a computer‐assisted analysis using the MetaboScape software suite for the determination of new metabolites .…”

“…We recently reported on the MatB_Sc from S. cinnamonensis . A biocatalytic assay with purified MatB_Sc was performed to further explore its substrate scope.…”

“…Substrate acceptance was monitored by direct product detection with HRMS/MS 2 . HRMS data for the main H + ‐adduct are shown in the table enclosed, and further adducts, characteristic fragmentation pattern, isotopic pattern and MS 2 data are shown in the Supporting Information (Tables S1 and S2, Figures S1 and S2); 4 – 6 and 11 – 12 were detected in an earlier report on this enzyme . Additional data were obtained for 8 – 16 in Michaelis–Menten kinetics (Figure ).…”

“…While direct supplementation of CoA‐activated malonate derivatives is unsuitable due to its limited membrane permeability and synthetic accessibility, N ‐acetylcysteamine esters (SNAC esters), acting as a CoA‐mimic, have been successfully used in feeding experiments . However, significant synthetic effort ahead of the fermentation, toxicity in fermentations and inferior activation compared to CoA‐thioesters have limited their application …”

“…We recently introduced a new MatB‐type enzyme from the monensin producer S. cinnamonensis and demonstrated its in vivo activity on a small number of different malonic acid derivatives, outcompeting artificial SNAC activation …”