The solution conformation of the peptide antibiotic thiostrepton. A 1H NMR study.

Abstract

The majority of the 84 protons in the 'H NMR spectrum of thiostrepton at 300 MHz were unambiguously assigned on the basis of double resonance experiments under different conditions of solvent, temperature and 214-exchange by comparison with the known crystal structure determined by ANDERSON et al.1) Evidence is presented to suggest that the side chain, the nature of which remained undefined on X-ray analysis, is comprised of two dehydroalanine residues which supports the conclusions reached by TORT et al.') on the basis of 1"C NMR spectroscopy. These two residues are missing in thiostrepton A;, a minor artifact. All available 'H NMR evidence suggests thiostrepton to have a similar conformation in deuterochloroform solution to that found in the crystal form.In our structural work on the thiopeptins3), which is the subject of a companion paper, we became heavily dependent on the 'H NMR assignments in the structurally related antibiotic thiostrepton, the structure of which was largely determined by X-ray crystallography by ANDERSON et al.1). Some doubt remained about the nature of the side chain which was subsequently shown to consist of two dehydroalanine (Deala) residues on the basis of 11C NMR spectroscopy2). We have therefore undertaken an extensive 'H NMR investigation of thiostrepton at 300 MHz under a wide variety of conditions which has resulted in unequivocal assignments of the majority of resonances. The 'H NMR evidence was found to be consistent with its X-ray structure and unique conformation, as well as confirming the nature of the side chain. Some of our findings were reported independently by ToRi et al.'). ExperimentalA sample of thiostrepton was obtained from the Squibb Institute. On standing it slowly decomposed to a minor product of higher Rf on silica gel TLC and was designated thiostrepton A2. It was easily separated from thiostrepton (for convenience assigned the A, component) by preparative TLC on silica gel GF (254) plates (Analtech) using CH, Cl, (1: 19).'H NMR spectra of thiostreptons A l and A, were obtained using a Varian SC-300 MHz spectrometer in the Fourier Transform mode using acquisition times of I and 2 seconds with sweep width of 4,000 Hz. Concentrations were in the range 3.5 -5.0 mg/0.4 ml and chemical shifts are quoted in 8, ppm downfield of TMS. Crystal coordinates for two althiomycin derivatives were obtained from published data5,6) from which torsional angles and internuclear bond distances were readily computed. Hydrogen positions in thiostrepton were calculated from the crystal coordinates1) assuming standard distances and angles.

Isolation and Structure Elucidation of Thiazomycin

“…However, key attributes of this class of natural products, such as highly potent activity and new modes of action, prompted us to study them further. Our research laboratories studied thiostrepton in the early eighties without making much progress in terms of development [8,9]. Significant advances in biology and chemistry have taken place since then.…”

Biosynthesis of the Thiopeptins and Identification of an F420H2-Dependent Dehydropiperidine Reductase

“…On the basis of the previously determined structure of 1 , we propose that the newly introduced stereocenter in 7 presents the same absolute configuration as is observed in the other series a thiopeptide metabolites 2 and 3 . 5,24 The piperidine ring in 7 , characteristic of a series a thiopeptide, suggests that TpnL reduces a precursor dehydropiperidine.…”

Form and Function in Cyclic Peptide Natural Products: A Pharmacokinetic Perspective

“…However, another group reported recently that the down-regulation of FOXM1 in malignant mesothelioma cells is not related to proteasome inhibition, but rather correlates with the ability of thiostrepton to disable the mitochondrial anti-oxidant pathway by covalently dimer- Despite the conflicting hypotheses regarding the MOA of thiostrepton, its target(s) is/are likely intracellular. The Xray [98] and solution NMR [99] structures of thiostrepton show some intramolecular hydrogen bonding, but also a significant number of solvent-exposed polar groups, suggesting that thiostrepton is unlikely to traverse membranes by a simple passive diffusion-type mechanism. However, the lack of cationic residues suggests its mechanism of membrane penetration is distinct from the pore-forming and membranedisrupting mechanisms shared by the classical amphiphilic antimicrobial peptides.…”