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The high resolution structure of tyrocidine A reveals an amphipathic dimer.

ABSTRACT: Tyrocidine A, one of the first antibiotics ever to be discovered, is a cyclic decapeptide that binds to membranes of target bacteria, disrupting their integrity. It is active against a broad spectrum of Gram-positive organisms, and has recently engendered interest as a potential scaffold for the development of new drugs to combat antibiotic-resistant pathogens. We present here the X-ray crystal structure of tyrocidine A at a resolution of 0.95Å. The structure reveals that tyrocidine forms an intimate and highly amphipathic homodimer made up of four beta strands that associate into a single, highly curved antiparallel beta sheet. We used surface plasmon resonance and potassium efflux assays to demonstrate that tyrocidine binds tightly to mimetics of bacterial membranes with an apparent dissociation constant (K(D)) of 10 ?M, and efficiently permeabilizes bacterial cells at concentrations equal to and below the K(D). Using variant forms of tyrocidine in which the fluorescent probe p-cyano-phenylalanine had been inserted on either the polar or apolar face of the molecule, we performed fluorescence quenching experiments, using both water-soluble and membrane-embedded quenchers. The quenching results, together with the structure, strongly support a membrane association model in which the convex, apolar face of tyrocidine's beta sheet is oriented toward the membrane interior, while the concave, polar face is presented to the aqueous phase.

SUBMITTER: Loll PJ

PROVIDER: S-EPMC3980540 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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The high resolution structure of tyrocidine A reveals an amphipathic dimer.

Loll Patrick J PJ Upton Elizabeth C EC Nahoum Virginie V Economou Nicoleta J NJ Cocklin Simon S

Biochimica et biophysica acta 20140211 5

Tyrocidine A, one of the first antibiotics ever to be discovered, is a cyclic decapeptide that binds to membranes of target bacteria, disrupting their integrity. It is active against a broad spectrum of Gram-positive organisms, and has recently engendered interest as a potential scaffold for the development of new drugs to combat antibiotic-resistant pathogens. We present here the X-ray crystal structure of tyrocidine A at a resolution of 0.95Å. The structure reveals that tyrocidine forms an int ...[more]

PMID: 24530898

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Project description:In bacteria, the mechanisms used to repair DNA lesions during genome replication include homologous recombination between sister chromosomes. This can lead to the formation of chromosome dimers if an odd number of crossover events occurs. The dimers must be resolved before cell separation to ensure genomic stability and cell viability. Dimer resolution is achieved by the broadly conserved dif/Xer system, which catalyzes one additional crossover event immediately prior to cell separation. While dif/Xer systems have been characterized or predicted in the vast majority of proteobacteria, no homologs to dif or xer have been identified in the order Legionellales. Here, we report the discovery of a distinct single-recombinase dif/Xer system in the intracellular pathogen Legionella pneumophila. The dif site was uncovered by our analysis of Legionella mobile element-1 (LME-1), which harbors a dif site mimic and integrates into the L. pneumophila genome via site-specific recombination. We demonstrate that lpg1867 (here named xerL) encodes a tyrosine recombinase that is necessary and sufficient for catalyzing recombination at the dif site and that deletion of dif or xerL causes filamentation along with extracellular and intracellular growth defects. We show that the dif/XerL system is present throughout Legionellales and that Coxiella burnetii XerL and its cognate dif site can functionally substitute for the native system in L. pneumophila. Finally, we describe an unexpected link between C. burnetii dif/Xer and the maintenance of its virulence plasmids. IMPORTANCE The maintenance of circular chromosomes depends on the ability to resolve aberrant chromosome dimers after they form. In most proteobacteria, broadly conserved Xer recombinases catalyze single crossovers at short, species-specific dif sites located near the replication terminus. Chromosomal dimerization leads to the formation of two copies of dif within the same molecule, leading to rapid site-specific recombination and conversion back into chromosome monomers. The apparent absence of chromosome dimer resolution mechanisms in Legionellales has been a mystery to date. By studying a phage-like mobile genetic element, LME-1, we have identified a previously unknown single-recombinase dif/Xer system that is not only widespread across Legionellales but whose activity is linked to virulence in two important human pathogens.

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The high resolution structure of tyrocidine A reveals an amphipathic dimer.

Publications

The high resolution structure of tyrocidine A reveals an amphipathic dimer.

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