Searching for Glycosylated Natural Products in Actinomycetes and Identification of Novel Macrolactams and Angucyclines

Abstract

Many bioactive natural products are glycosylated compounds in which the sugar components usually participate in interaction and molecular recognition of the cellular target. Therefore, the presence of sugar moieties is important, in some cases essential, for bioactivity. Searching for novel glycosylated bioactive compounds is an important aim in the field of the research for natural products from actinomycetes. A great majority of these sugar moieties belong to the 6-deoxyhexoses and share two common biosynthetic steps catalyzed by a NDP-D-glucose synthase (GS) and a NDP-D-glucose 4,6-dehydratase (DH). Based on this fact, seventy one Streptomyces strains isolated from the integument of ants of the Tribe Attini were screened for the presence of biosynthetic gene clusters (BGCs) for glycosylated compounds. Total DNAs were analyzed by PCR amplification using oligo primers for GSs and DHs and also for a NDP-D-glucose-2,3-dehydratases. Amplicons were used in gene disruption experiments to generate non-producing mutants in the corresponding clusters. Eleven mutants were obtained and comparative dereplication analyses between the wild type strains and the corresponding mutants allowed in some cases the identification of the compound coded by the corresponding cluster (lobophorins, vicenistatin, chromomycins and benzanthrins) and that of two novel macrolactams (sipanmycin A and B). Several strains did not show UPLC differential peaks between the wild type strain and mutant profiles. However, after genome sequencing of these strains, the activation of the expression of two clusters was achieved by using nutritional and genetic approaches leading to the identification of compounds of the cervimycins family and two novel members of the warkmycins family. Our work defines a useful strategy for the identification new glycosylated compounds by a combination of genome mining, gene inactivation experiments and the activation of silent biosynthetic clusters in Streptomyces strains.

Cooperative Involvement of Glycosyltransferases in the Transfer of Amino Sugars during the Biosynthesis of the Macrolactam Sipanmycin by Streptomyces sp. Strain CS149

“…strain ML694-90F31 cluster involved in incednine (idn) biosynthesis (22) (Table 1). Furthermore, three genes of the cluster involved in amino sugar biosynthesis (sipS6, sipS7, and sipS10) contained DNA sequences that were previously used to generate sipanmycin-nonproducing mutants in this strain (13). In addition, a cluster very similar to the sip cluster is also present in Streptomyces sp.…”

“…Streptomyces sp. CS149, a sipanmycin producer, has been previously reported (13). Medium A (MA) (53) was used for sporulation.…”

“…We have recently applied a targeted screening in the search for the presence of glycosylated bioactive natural products in a collection of Streptomyces strains isolated from leaf-cutting ants (13). By combining PCR-based screening, mass spectrometry (MS) dereplication, and generation of mutants, we identified two novel macrolactams, sipanmycins A and B ( Fig.…”

Identification and characterization of the first β-1,3-d-xylosidase from a gram-positive bacterium, Streptomyces sp. SWU10

“…WP_087792600.1: 95 %) of Streptomyces sp. CS159[29], and xylosidase/arabinofuranosidase (GenBank accession no. EHN79169.1: 95 %) of Streptomyces coelicoflavus ZG0656[30]…”

α-d-Glucopyranosyl-(1→2)-[6-O-(l-tryptophanyl)-β-d-fructofuranoside]

“…Glycosylated natural products are defined as natural compounds where the carbohydrate part (glycone), either monomers or oligomers of different structures, is covalently bound to another chemical structure (aglycone). The aglycone encompasses natural product scaffolds such as terpenes, steroids, alkaloids, ribosomal and non-ribosomal peptides, modified amino acids, polyketides, macrolides, flavonoids, polyenes, coumarins, anthracyclines, iridoids and lignans [13][14][15][16]. Glycosides display Molbank 2019, 2019, M1066 2 of 11 very broad bioactivities because, part of their structures are composed of one or more stereocenter laden sugar portions with numerous hydroxyls and other functionalities that confer increased drug water solubility, increased bioavailability, decreased toxicity, effective drug targeting, bioactivity, strong molecular targeting and organism specificity while the non-sugar portion or aglycones, represent molecules that also have many bioactivities [17].…”