Architecture of Microcin B17 Synthetase: An Octameric Protein Complex Converting a Ribosomally Synthesized Peptide into a DNA Gyrase Poison.

The introduction of azole heterocycles into a peptide backbone is the principal step in the biosynthesis of numerous compounds with therapeutic potential. One of them is microcin B17, a bacterial topoisomerase inhibitor whose activity depends on the conversion of selected serine and cysteine residues of the precursor peptide to oxazoles and thiazoles by the McbBCD synthetase complex. Crystal structures of McbBCD reveal an octameric B4C2D2 complex with two bound substrate peptides. Each McbB dimer clamps the N-terminal recognition sequence, while the C-terminal heterocycle of the modified peptide is trapped in the active site of McbC. The McbD and McbC active sites are distant from each other, which necessitates alternate shuttling of the peptide substrate between them, while remaining tethered to the McbB dimer. An atomic-level view of the azole synthetase is a starting point for deeper understanding and control of biosynthesis of a large group of ribosomally synthesized natural products.

Expression of genes involved in carbohydrate-lipid metabolism in muscle and fat tissues in the initial stage of adult-age obesity in fed and fasted mice.

C57Bl mice exhibit impaired glucose metabolism by the late adult age under standard living conditions. The aim of this study was to evaluate white adipose tissue (WAT), brown adipose tissue (BAT), and skeletal muscle expression of genes involved in carbohydrate-lipid metabolism at postpubertal stages preceding the late adult age in C57Bl mice. Muscle mRNA levels of uncoupling protein 3 (Ucp3) and carnitine palmitoyltransferase 1 (Cpt1) (indicators of FFA oxidation), WAT mRNA levels of hormone-sensitive lipase (Lipe) and lipoprotein lipase (Lpl) (indicators of lipolysis and lipogenesis), muscle and WAT mRNA levels of the type 4 glucose transporter Slc2a4 (indicators of insulin-dependent glucose uptake), and BAT mRNA levels of uncoupling protein 1 (Ucp1) (indicator of thermogenesis) were measured in fed and 16 h-fasted mice in three age groups: 10-week-old (young), 15-week-old (early adult), and 30-week-old (late adult). Weight gain from young to early adult age was not accompanied by changes in WAT and BAT indexes and biochemical blood parameters. Weight gain from early to late adult age was accompanied by increased WAT and BAT indexes and decreased glucose tolerance. Muscle Ucp3 and Cpt1 mRNA levels and WAT Lipe and Slc2a4 mRNA levels increased from young to early adult age and then sharply decreased by the late adult age. Moreover, BAT Ucp1 mRNA level decreased in the late adult age. Fasting failed to increase muscle Cpt1 mRNA levels in late adult mice. These transcriptional changes could contribute to impaired glucose metabolism and the onset of obesity in late adult mice during normal development.

The Product of Yersinia pseudotuberculosis mcc Operon Is a Peptide-Cytidine Antibiotic Activated Inside Producing Cells by the TldD/E Protease.

Microcin C is a heptapeptide-adenylate antibiotic produced by some strains of Escherichia coli. Its peptide part is responsible for facilitated transport inside sensitive cells where it is proteolyzed with release of a toxic warhead-a nonhydrolyzable aspartamidyl-adenylate, which inhibits aspartyl-tRNA synthetase. Recently, a microcin C homologue from Bacillus amyloliquefaciens containing a longer peptide part modified with carboxymethyl-cytosine instead of adenosine was described, but no biological activity of this compound was revealed. Here, we characterize modified peptide-cytidylate from Yersinia pseudotuberculosis. As reported for B. amyloliquefaciens homologue, the initially synthesized compound contains a long peptide that is biologically inactive. This compound is subjected to endoproteolytic processing inside producing cells by the evolutionary conserved TldD/E protease. As a result, an 11-amino acid long peptide with C-terminal modified cytosine residue is produced. This compound is exported outside the producing cell and is bioactive, inhibiting sensitive cells in the same way as E. coli microcin C. Proteolytic processing inside producing cells is a novel strategy of peptide-nucleotide antibiotics biosynthesis that may help control production levels and avoid toxicity to the producer.

Peptide-nucleotide antibiotic Microcin C is a potent inducer of stringent response and persistence in both sensitive and producing cells.

Microcin C (McC) is a peptide-nucleotide antibiotic that inhibits aspartyl-tRNA synthetase. Here, we show that McC is a strong inducer of persistence in Escherichia coli. Persistence induced by McC is mediated by (p)ppGpp and requires chromosomally encoded toxin-antitoxin modules. McC-producing cells have increased persistence levels due to a combined effect of McC imported from the cultured medium and intracellularly synthesized antibiotic. McC-producing cells also induce persistence in sensitive cells during co-cultivation, underscoring complex interactions in bacterial communities where an antagonistic compound produced by one community member can benefit other members by increasing their ability to withstand antibiotics.