Solid-state NMR measurements performed on intact whole cells of Staphylococcus aureus labeled selectively in vivo have established that des-N-methylleucyl oritavancin (which has antimicrobial activity) binds to the cell-wall peptidoglycan, even though removal of the terminal N-methylleucyl residue destroys the D-Ala-D-Ala binding pocket. By contrast, the des-N-methylleucyl form of vancomycin (which has no antimicrobial activity) does not bind to the cell wall. Solid-state NMR has also determined that oritavancin and vancomycin are comparable inhibitors of transglycosylation, but that oritavancin is a more potent inhibitor of transpeptidation. This combination of effects on cell-wall binding and biosynthesis is interpreted in terms of a recent proposal that oritavancin-like glycopeptides have two cell-wall binding sites: the well-known peptidoglycan D-Ala-D-Ala pentapeptide stem terminus and the pentaglycyl bridging segment. The resulting dual mode of action provides a structural framework for coordinated cell-wall assembly that accounts for the enhanced potency of oritavancin and oritavancin-like analogues against vancomycin-resistant organisms.

The increasing frequency of Enterococcus faecium isolates with multidrug resistance is a serious clinical problem given the severely limited number of therapeutic options available to treat these infections. Oritavancin is a promising new alternative in clinical development that has potent antimicrobial activity against both staphylococcal and enterococcal vancomycin-resistant pathogens. Using solid-state NMR to detect changes in the cell-wall structure and peptidoglycan precursors of whole cells after antibiotic-induced stress, we report that vancomycin and oritavancin have different modes of action in E. faecium. Our results show the accumulation of peptidoglycan precursors after vancomycin treatment, consistent with transglycosylase inhibition, but no measurable difference in cross-linking. In contrast, after oritavancin exposure, we do not observe the accumulation of peptidoglycan precursors. Instead, the number of cross-links is significantly reduced, showing that oritavancin primarily inhibits transpeptidation. We propose that the activity of oritavancin is the result of a secondary-binding interaction with the E. faecium peptidoglycan. The hypothesis is supported by results from 13 C{ 19 F} REDOR experiments on whole cells enriched with L-[1-13 C] lysine and complexed with desleucyl [ 19 F]oritavancin. These experiments establish that an oritavancin derivative with a damaged D-Ala-D-Ala binding pocket still binds to E. faecium peptidoglycan. The 13 C{ 19 F} REDOR dephasing maximum indicates that the secondary-binding site of oritavancin is specific to nascent and template peptidoglycan. We conclude that the inhibition of transpeptidation by oritavancin in E. faecium is the result of the large number of secondary-binding sites relative to the number of primary-binding sites.

A synthetic receptor is described with the appropriate shape and size for alkylated trimethylammonium ions such as choline and carnitine. The structure features a deep, concave binding site, lined with aromatic walls that provide cation-pi interactions between host and guest. Molecular mechanic calculations suggest that the host's shape is maintained through intermolecular hydrogen bonding with DMSO solvent molecules. The cavity is too small to accommodate larger ions. Choline and carnitine are recognized and bound with high affinity even though no complementary charges are involved in the process.

Glycopeptides whose aminosugars have been modified by attachment of hydrophobic side chains are frequently active against vancomycin-resistant microorganisms. We have compared the conformations of six such fluorinated glycopeptides (with side chains of varying length) complexed to cell walls labeled with d-[1-13C]alanine, [1-13C]glycine, and l-[ε-15N]lysine in whole-cells of Staphylococcus aureus. The internuclear distances from 19F of the bound drug to the 13C and 15N labels of the peptidoglycan, and to the natural abundance 31P of lipid membranes and teichoic acids, were determined by rotational-echo double resonance NMR. The drugs did not dimerize, and their side chains did not form membrane anchors but instead became essential parts of secondary binding to pentaglycyl bridge segments of the cell-wall peptidoglycan.

Tetrabenzimidazole cavitands 4 were prepared by condensation of ortho esters with octaamino cavitand 3 in 70-80% yield. Molecular modeling predicted that no intramolecular hydrogen bonds are possible between the imidazole fragments in the vase conformation of 4. Instead, this conformation provides four perfect binding sites for hydroxyl-containing molecules through an N-H---O-H---N pattern. Such interactions provide the means for sealing the cavitand's cavity. Accordingly, dry compounds 4 are not soluble in dry CDCl3 but readily dissolve upon addition of small amounts of alcohols or by saturation of the solution with water. 1H NMR spectroscopy revealed that in these solutions molecules 4 adopt a vase conformation while 1D GOESY experiments revealed their monomeric nature. In water-saturated CDCl3, these cavitands 4 form kinetically stable 1:1 inclusion complexes with tetramethylphosphonium bromide and triethylammonium chloride in which the cation is incorporated into the pi-basic cavity. Thus, cavitands 4 are a novel class of open-ended molecular containers capable of the formation of highly kinetically stable complexes upon assistance by hydrogen-bonding water molecules.