Lipid II Research Articles

Structures of Streptococcus pyogenes class A sortase in complex with substrate and product mimics provide key details of target recognition

The cell wall is a critical extracellular barrier for bacteria and many other organisms. In bacteria, this structural layer consists of peptidoglycan, which maintains cell shape and structural integrity and provides a scaffold for displaying various protein factors. To attach proteins to the cell wall, Gram-positive bacteria utilize sortase enzymes, which are cysteine transpeptidases that recognize and cleave a specific sorting signal, followed by ligation of the sorting signal–containing protein to the peptidoglycan precursor lipid II (LII). This mechanism is the subject of considerable interest as a target for therapeutic intervention and as a tool for protein engineering, where sortases have enabled sortase-mediated ligation or sortagging strategies. Despite these uses, there remains an incomplete understanding of the stereochemistry of substrate recognition and ligation product formation. Here, we solved the first structures of sortase A from Streptococcus pyogenes bound to two substrate sequences, LPATA and LPATS. In addition, we synthesized a mimetic of the product of sortase-mediated ligation involving LII (LPAT-LII) and solved the complex structure in two ligand conformations. These structures were further used as the basis for molecular dynamics simulations to probe sortase A-ligand dynamics and to construct a model of the acyl–enzyme intermediate, thus providing a structural view of multiple key states in the catalytic mechanism. Overall, this structural information provides new insights into the recognition of the sortase substrate motif and LII ligation partner and will support the continued development of sortases for protein engineering applications.

Binding Modes of Teixobactin to Lipid II: Molecular Dynamics Study

Teixobactin (TXB) is a newly discovered antibiotic targeting the bacterial cell wall precursor Lipid II (LII). In the present work, four binding modes of TXB on LII were identified by a contact-map based clustering method. The highly flexible binary complex ensemble was generated by parallel tempering metadynamics simulation in a well-tempered ensemble (PTMetaD-WTE). In agreement with experimental findings, the pyrophosphate group and the attached first sugar subunit of LII are found to be the minimal motif for stable TXB binding. Three of the four binding modes involve the ring structure of TXB and have relatively higher binding affinities, indicating the importance of the ring motif of TXB in LII recognition. TXB-LII complexes with a ratio of 2:1 are also predicted with configurations such that the ring motif of two TXB molecules bound to the pyrophosphate-MurNAc moiety and the glutamic acid residue of one LII, respectively. Our findings disclose that the ring motif of TXB is critical to LII binding and novel antibiotics can be designed based on its mimetics.

Structural characterization of phosphatidylglycerol model membranes containing the antibiotic target lipid II molecule: a Raman microspectroscopy study

Fil: Sosa Morales, Marcelo Clemente. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Tucuman. Instituto de Quimica del Noroeste. Universidad Nacional de Tucuman. Facultad de Bioquimica, Quimica y Farmacia. Instituto de Quimica del Noroeste; Argentina

Correction: Turning Defense into Offense: Defensin Mimetics as Novel Antibiotics Targeting Lipid II

Correction: Turning Defense into Offense: Defensin Mimetics as Novel Antibiotics Targeting Lipid II

Structural dynamics of the cell wall precursor lipid II in the presence and absence of the lantibiotic nisin

Representing a physiological “Achilles' heel”, the cell wall precursor lipid II (LII) is a prime target for various classes of antibiotics. Over the years LII-binding agents have been recognized as promising candidates and templates in the search for new antibacterial compounds to complement or replace existing drugs. To elucidate the molecular structural basis underlying LII functional mechanism and to better understand if and how lantibiotic binding alters the molecular behavior of LII, we performed molecular dynamics (MD) simulations of phospholipid membrane-embedded LII in the absence and presence of the LII-binding lantibiotic nisin. In a series of 2×4 independent, unbiased 100ns MD simulations we sampled the conformational dynamics of nine LII as well as nine LII–nisin complexes embedded in an aqueous 150mM NaCl/POPC phospholipid membrane environment. We found that nisin binding to LII induces a reduction of LII mobility and flexibility, an outward shift of the LII pentapeptide, an inward movement of the LII disaccharide section, and an overall deeper insertion of the LII tail group into the membrane. The latter effect might indicate an initial step in adopting a stabilizing, scaffold-like structure in the process of nisin-induced membrane leakage. At the same time nisin conformation and LII interaction remain similar to the 1WCO LII–nisin NMR solution structure.

Getting Closer to the Real Bacterial Cell Wall Target: Biomolecular Interactions of Water‐Soluble Lipid II with Glycopeptide Antibiotics

A novel synthesized water-soluble variant of lipid II (LII) was used to evaluate the noncovalent interactions between a number of glycopeptide antibiotics and their receptor by bioaffinity electrospray ionization mass spectrometry (ESI-MS). The water-soluble variant of lipid II is an improved design, compared to the traditionally used tripeptide N,N'-diacetyl-L-lysyl-D-alanyl-D-alanine (KAA), of the target molecule on the bacterial cell wall. A representative group of glycopeptide antibiotics was selected for this study to evaluate the validity of the novel cell-wall-mimicking target LII. Structure-function relationships of various glycopeptide antibiotics were investigated by means of 1) bioaffinity mass spectrometry to evaluate solution-phase molecular interactions with both LII and KAA, 2) fluorescence leakage experiments to study the interactions with the membrane-embedded lipid II, and 3) minimum inhibitory concentrations against the indicator strain Micrococcus flavus. Our results with the novel LII molecule reveal that some antibiotics interact differently with KAA and LII. Additionally, our data cast doubt on the hypothesis that antibiotic selfdimerization assists in the in-vivo efficacy. Finally, the water-soluble lipid II proved to be a better model of the bacterial cell wall.