Biosynthesis of the Nuclear Factor of Activated T Cells Inhibitor NFAT-133 in Streptomyces pactum

Abstract

NFAT-133 is a Streptomyces-derived aromatic polyketide compound with immunosuppressive, antidiabetic, and antitrypanosomal activities. It inhibits transcription mediated by nuclear factor of activated T cells (NFAT), leading to the suppression of interleukin-2 expression and T cell proliferation. It also activates the AMPK pathway in L6 myotubes and increases glucose uptake. In addition to NFAT-133, a number of its congeners, e.g., panowamycins and benwamycins, have been identified. However, little is known about their modes of formation in the producing organisms. Through genome sequencing of Streptomyces pactum ATCC 27456, gene inactivation, and genetic complementation experiments, the biosynthetic gene cluster of NFAT-133 and its congeners has been identified. The cluster contains a highly disordered genetic organization of type I modular polyketide synthase genes with several genes that are necessary for the formation of the aromatic core unit and tailoring processes. In addition, a number of new analogs of NFAT-133 were isolated and their chemical structures elucidated. It is suggested that the heptaketide NFAT-133 is derived from an octaketide intermediate, TM-123. The current study shows yet another unusual biosynthetic pathway involving a noncanonical polyketide synthase assembly line to produce a group of small molecules with valuable bioactivities.

Antidiabetic profiling of veramycins, polyketides accessible by biosynthesis, chemical synthesis and precursor-directed modification

“…Its molecular formula was determined as C 21 H 26 O 5 based on HR-ESI-MS and its 1 H and 13 C NMR data showed complete identity with those reported for TM-123. 16 Veramycin C (18), obtained as a white powder was determined to have the molecular formula C 20 H 22 O 5 based on HR-ESI-MS, indicating ten degrees of unsaturation. A closer examination of the NMR data revealed the absence of a hydroxylated allylic residue at the aromatic ring in contrast to 13 and 17.…”

“…Optical rotations were determined as [α] 22 D = +114 (c = 1.02, MeOH), lit. 16 ] propionate were supplemented to fermentations. This revealed 13 to be biosynthesized by assembly of five methylmalonyl-and three malonyl-CoA units (Fig.…”

“…15 While the preparation of our manuscript about investigations on the isolation, characterization, biosynthesis and total synthesis of NFAT-133 and its congeners, the BGC corresponding to NFAT-133 production in Streptomyces pactum was published along with the disclosure of further congeners (Chart 1). 16 The 2D structure of NFAT-133 was determined in 1995 11 but only in 2016, a first proposal of the absolute configuration of NFAT-133 was published based on data obtained from NOESY experiments, J-based configuration analysis ( JBCA) and advanced Mosher's analysis. 17 For the contiguous three stereogenic centers the absolute configurations were proposed as 10R, 11R and 12S, respectively, representing a syn,anti (threo, erythro) stereotriad (or stereotriad "B" according to the Hoffmann nomenclature).…”

“…After bioassay and UPLC-HRMS guided fractionation, the antibacterial activity of the extract against E. coli was assigned to the presence of pactamycin. After re-fermentation of the strain, the recently disclosed 16 TM-123 (13) as well as 7 new polyketide natural products 17-23 were isolated from larger batches of the extract (Chart 2). A closely related congener termed veramycin A ( 17) with an α-pyrone motif particularly gained our attraction, since it showed a comparable effect to NFAT-133 on glucose uptake in L6 rat skeletal myotubes.…”

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“…Genome sequencing of Streptomyces pactum has identified the biosynthetic gene cluster of NFAT-133 27 , an aromatic polyketide with immunosuppressive, anti-diabetic and anti-trypanosomal activities. 24 Analysis of the biosynthetic pathway, which involves a non-canonical PKS assembly line, suggests the formation of heptaketide 27 by a novel process involving the lactone ring cleavage of octaketide TM-123 28 .…”