Structural variations of the cell wall precursor lipid II in Gram-positive bacteria — Impact on binding and efficacy of antimicrobial peptides

ABSTRACTResistance against cell wall-active antimicrobial peptides in bacteria is often mediated by transporters. In low GC-content Gram-positive bacteria, a wide-spread type of such transporters are the BceAB-like systems, which frequently provide a high level of resistance against peptide antibiotics that target intermediates of the lipid II cycle of cell wall synthesis. How a transporter can offer protection from drugs that are active on the cell surface, however, has presented researchers with a conundrum. Multiple theories have been discussed, ranging from removal of the peptides from the membrane, internalisation of the drug for degradation, to removal of the cellular target rather than the drug itself. To resolve this much-debated question, we here investigated the mode of action of the transporter BceAB of Bacillus subtilis. We show that it does not inactivate or import its substrate antibiotic bacitracin. Moreover, we present evidence that the critical factor driving transport activity is not the drug itself, but instead the concentration of drug-target complexes in the cell. Our results, together with previously reported findings, lead us to propose that BceAB-type transporters act by transiently freeing lipid II cycle intermediates from the inhibitory grip of antimicrobial peptides, and thus provide resistance through target protection of cell wall synthesis. Target protection has so far only been reported for resistance against antibiotics with intracellular targets, such as the ribosome. However, this mechanism offers a plausible explanation for the use of transporters as resistance determinants against cell wall-active antibiotics in Gram-positive bacteria where cell wall synthesis lacks the additional protection of an outer membrane.

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BceAB-Type Antibiotic Resistance Transporters Appear To Act by Target Protection of Cell Wall Synthesis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02241-19 ◽

2019 ◽

Vol 64 (3) ◽

Cited By ~ 6

Author(s):

Carolin M. Kobras ◽

Hannah Piepenbreier ◽

Jennifer Emenegger ◽

Andre Sim ◽

Georg Fritz ◽

...

Keyword(s):

Cell Wall ◽

Antimicrobial Peptides ◽

Gc Content ◽

Critical Factor ◽

Peptide Antibiotics ◽

Cell Wall Synthesis ◽

Gram Positive ◽

Content Type ◽

Gram Positive Bacteria ◽

Lipid Ii

ABSTRACT Resistance against cell wall-active antimicrobial peptides in bacteria is often mediated by transporters. In low-GC-content Gram-positive bacteria, a common type of such transporters is BceAB-like systems, which frequently provide high-level resistance against peptide antibiotics that target intermediates of the lipid II cycle of cell wall synthesis. How a transporter can offer protection from drugs that are active on the cell surface, however, has presented researchers with a conundrum. Multiple theories have been discussed, ranging from removal of the peptides from the membrane and internalization of the drug for degradation to removal of the cellular target rather than the drug itself. To resolve this much-debated question, we here investigated the mode of action of the transporter BceAB of Bacillus subtilis. We show that it does not inactivate or import its substrate antibiotic bacitracin. Moreover, we present evidence that the critical factor driving transport activity is not the drug itself but instead the concentration of drug-target complexes in the cell. Our results, together with previously reported findings, lead us to propose that BceAB-type transporters act by transiently freeing lipid II cycle intermediates from the inhibitory grip of antimicrobial peptides and thus provide resistance through target protection of cell wall synthesis. Target protection has so far only been reported for resistance against antibiotics with intracellular targets, such as the ribosome. However, this mechanism offers a plausible explanation for the use of transporters as resistance determinants against cell wall-active antibiotics in Gram-positive bacteria where cell wall synthesis lacks the additional protection of an outer membrane.

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Structural Variations of the Cell Wall Precursor Lipid II and Their Influence on Binding and Activity of the Lipoglycopeptide Antibiotic Oritavancin

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02663-14 ◽

2014 ◽

Vol 59 (2) ◽

pp. 772-781 ◽

Cited By ~ 27

Author(s):

Daniela Münch ◽

Ina Engels ◽

Anna Müller ◽

Katrin Reder-Christ ◽

Hildegard Falkenstein-Paul ◽

...

Keyword(s):

Cell Wall ◽

Binding Affinity ◽

Specific Binding ◽

Intramolecular Interactions ◽

Structural Variations ◽

Gram Positive ◽

Content Type ◽

Lipid Ii ◽

Vancomycin Resistant ◽

The Impact

ABSTRACTOritavancin is a semisynthetic derivative of the glycopeptide antibiotic chloroeremomycin with activity against Gram-positive pathogens, including vancomycin-resistant staphylococci and enterococci. Compared to vancomycin, oritavancin is characterized by the presence of two additional residues, a hydrophobic 4′-chlorobiphenyl methyl moiety and a 4-epi-vancosamine substituent, which is also present in chloroeremomycin. Here, we show that oritavancin and its des-N-methylleucyl variant (des-oritavancin) effectively inhibit lipid I- and lipid II-consuming peptidoglycan biosynthesis reactionsin vitro. In contrast to that for vancomycin, the binding affinity of oritavancin to the cell wall precursor lipid II appears to involve, in addition to thed-Ala-d-Ala terminus, other species-specific binding sites of the lipid II molecule, i.e., the crossbridge andd-isoglutamine in position 2 of the lipid II stem peptide, both characteristic for a number of Gram-positive pathogens, including staphylococci and enterococci. Using purified lipid II and modified lipid II variants, we studied the impact of these modifications on the binding of oritavancin and compared it to those of vancomycin, chloroeremomycin, and des-oritavancin. Analysis of the binding parameters revealed that additional intramolecular interactions of oritavancin with the peptidoglycan precursor appear to compensate for the loss of a crucial hydrogen bond in vancomycin-resistant strains, resulting in enhanced binding affinity. Augmenting previous findings, we show that amidation of the lipid II stem peptide predominantly accounts for the increased binding of oritavancin to the modified intermediates ending ind-Ala-d-Lac. Corroborating our conclusions, we further provide biochemical evidence for the phenomenon of the antagonistic effects ofmecAandvanAresistance determinants inStaphylococcus aureus, thus partially explaining the low frequency of methicillin-resistantS. aureus(MRSA) acquiring high-level vancomycin resistance.

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Mechanism of Action of the Mannopeptimycins, a Novel Class of Glycopeptide Antibiotics Active against Vancomycin-Resistant Gram-Positive Bacteria

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.3.728-738.2004 ◽

2004 ◽

Vol 48 (3) ◽

pp. 728-738 ◽

Cited By ~ 70

Author(s):

Alexey Ruzin ◽

Guy Singh ◽

Anatoly Severin ◽

Youjun Yang ◽

Russell G. Dushin ◽

...

Keyword(s):

Cell Wall ◽

Lipoteichoic Acid ◽

Antimicrobial Activities ◽

Cell Wall Biosynthesis ◽

Glycopeptide Antibiotics ◽

Gram Positive ◽

Gram Positive Bacteria ◽

Lipid Ii ◽

Membrane Bound ◽

Vancomycin Resistant

ABSTRACT The naturally occurring mannopeptimycins (formerly AC98-1 through AC98-5) are a novel class of glycopeptide antibiotics that are active against a wide variety of gram-positive bacteria. The structures of the mannopeptimycins suggested that they might act by targeting cell wall biosynthesis, similar to other known glycopeptide antibiotics; but the fact that the mannopeptimycins retain activity against vancomycin-resistant organisms suggested that they might have a unique mode of action. By using a radioactive mannopeptimycin derivative bearing a photoactivation ligand, it was shown that mannopeptimycins interact with the membrane-bound cell wall precursor lipid II [C55-MurNAc-(peptide)-GlcNAc] and that this interaction is different from the binding of other lipid II-binding antibiotics such as vancomycin and mersacidin. The antimicrobial activities of several mannopeptimycin derivatives correlated with their affinities toward lipid II, suggesting that the inhibition of cell wall biosynthesis was primarily through lipid II binding. In addition, it was shown that mannopeptimycins bind to lipoteichoic acid in a rather nonspecific interaction, which might facilitate the accumulation of antibiotic on the bacterial cell surface.

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The dlt operon confers resistance to cationic antimicrobial peptides in Clostridium difficile

Microbiology ◽

10.1099/mic.0.045997-0 ◽

2011 ◽

Vol 157 (5) ◽

pp. 1457-1465 ◽

Cited By ~ 91

Author(s):

Shonna M. McBride ◽

Abraham L. Sonenshein

Keyword(s):

Cell Wall ◽

Clostridium Difficile ◽

Antimicrobial Peptides ◽

Positive Charge ◽

Polymyxin B ◽

Gram Positive ◽

Cationic Antimicrobial Peptides ◽

Teichoic Acids ◽

Gram Positive Bacteria ◽

Net Positive Charge

The dlt operon in Gram-positive bacteria encodes proteins that are necessary for the addition of d-alanine to teichoic acids of the cell wall. The addition of d-alanine to the cell wall results in a net positive charge on the bacterial cell surface and, as a consequence, can decrease the effectiveness of antimicrobials, such as cationic antimicrobial peptides (CAMPs). Although the roles of the dlt genes have been studied for some Gram-positive organisms, the arrangement of these genes in Clostridium difficile and the life cycle of the bacterium in the host are markedly different from those of other pathogens. In the current work, we determined the contribution of the putative C. difficile dlt operon to CAMP resistance. Our data indicate that the dlt operon is necessary for full resistance of C. difficile to nisin, gallidermin, polymyxin B and vancomycin. We propose that the d-alanylation of teichoic acids provides protection against antimicrobial peptides that may be essential for growth of C. difficile in the host.

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Increasing the Antimicrobial Activity of Nisin-Based Lantibiotics against Gram-Negative Pathogens

Applied and Environmental Microbiology ◽

10.1128/aem.00052-18 ◽

2018 ◽

Vol 84 (12) ◽

Cited By ~ 31

Author(s):

Qian Li ◽

Manuel Montalban-Lopez ◽

Oscar P. Kuipers

Keyword(s):

Antimicrobial Peptides ◽

Outer Membrane ◽

Rational Design ◽

Bacterial Species ◽

Gram Negative Bacteria ◽

Gram Positive ◽

Gram Negative ◽

Content Type ◽

Gram Positive Bacteria ◽

Lipid Ii

ABSTRACTLantibiotics are ribosomally synthesized and posttranslationally modified antimicrobial compounds containing lanthionine and methyl-lanthionine residues. Nisin, one of the most extensively studied and used lantibiotics, has been shown to display very potent activity against Gram-positive bacteria, and stable resistance is rarely observed. By binding to lipid II and forming pores in the membrane, nisin can cause the efflux of cellular constituents and inhibit cell wall biosynthesis. However, the activity of nisin against Gram-negative bacteria is much lower than that against Gram-positive bacteria, mainly because lipid II is located at the inner membrane, and the rather impermeable outer membrane in Gram-negative bacteria prevents nisin from reaching lipid II. Thus, if the outer membrane-traversing efficiency of nisin could be increased, the activity against Gram-negative bacteria could, in principle, be enhanced. In this work, several relatively short peptides with activity against Gram-negative bacteria were selected from literature data to be fused as tails to the C terminus of either full or truncated nisin species. Among these, we found that one of three tails (tail 2 [T2; DKYLPRPRPV], T6 [NGVQPKY], and T8 [KIAKVALKAL]) attached to a part of nisin displayed improved activity against Gram-negative microorganisms. Next, we rationally designed and reengineered the most promising fusion peptides. Several mutants whose activity significantly outperformed that of nisin against Gram-negative pathogens were obtained. The activity of the tail 16 mutant 2 (T16m2) construct against several important Gram-negative pathogens (i.e.,Escherichia coli,Klebsiella pneumoniae,Acinetobacter baumannii,Pseudomonas aeruginosa,Enterobacter aerogenes) was increased 4- to 12-fold compared to that of nisin. This study indicates that the rational design of nisin can selectively and significantly improve its outer membrane-permeating capacity as well as its activity against Gram-negative pathogens.IMPORTANCELantibiotics are antimicrobial peptides that are highly active against Gram-positive bacteria but that have relatively poor activity against most Gram-negative bacteria. Here, we modified the model lantibiotic nisin by fusing parts of it to antimicrobial peptides with known activity against Gram-negative bacteria. The appropriate selection of peptidic moieties that could be attached to (parts of) nisin could lead to a significant increase in its inhibitory activity against Gram-negative bacteria. Using this strategy, hybrids that outperformed nisin by displaying 4- to 12-fold higher levels of activity against relevant Gram-negative bacterial species were produced. This study shows the power of modified peptide engineering to alter target specificity in a desired direction.

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Influence of Cell Wall Composition on the Fossilisation of Bacteria and the Implications for the Search for Early Life Forms

International Astronomical Union Colloquium ◽

10.1017/s0252921100015025 ◽

1997 ◽

Vol 161 ◽

pp. 491-504 ◽

Cited By ~ 3

Author(s):

Frances Westall

Keyword(s):

Cell Wall ◽

Life Forms ◽

Cation Binding ◽

Positive Bacterium ◽

Gram Positive ◽

Gram Positive Bacteria ◽

Transmission Electron ◽

Hydrothermal Sediments ◽

Identification Of Bacteria ◽

The Difference

AbstractThe oldest cell-like structures on Earth are preserved in silicified lagoonal, shallow sea or hydrothermal sediments, such as some Archean formations in Western Australia and South Africa. Previous studies concentrated on the search for organic fossils in Archean rocks. Observations of silicified bacteria (as silica minerals) are scarce for both the Precambrian and the Phanerozoic, but reports of mineral bacteria finds, in general, are increasing. The problems associated with the identification of authentic fossil bacteria and, if possible, closer identification of bacteria type can, in part, be overcome by experimental fossilisation studies. These have shown that not all bacteria fossilise in the same way and, indeed, some seem to be very resistent to fossilisation. This paper deals with a transmission electron microscope investigation of the silicification of four species of bacteria commonly found in the environment. The Gram positiveBacillus laterosporusand its spore produced a robust, durable crust upon silicification, whereas the Gram negativePseudomonas fluorescens, Ps. vesicularis, andPs. acidovoranspresented delicately preserved walls. The greater amount of peptidoglycan, containing abundant metal cation binding sites, in the cell wall of the Gram positive bacterium, probably accounts for the difference in the mode of fossilisation. The Gram positive bacteria are, therefore, probably most likely to be preserved in the terrestrial and extraterrestrial rock record.

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Microanatomy of the Periplasm of Bacillus Licheniformis

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100065365 ◽

1971 ◽

Vol 29 ◽

pp. 262-263

Author(s):

B.K. Ghosh

Keyword(s):

Cell Wall ◽

Plasma Membrane ◽

Bacillus Licheniformis ◽

External Environment ◽

Permeability Barrier ◽

Periplasmic Space ◽

Modified Technique ◽

Gram Positive ◽

Gram Negative ◽

Gram Positive Bacteria

Periplasm of bacteria is the space outside the permeability barrier of plasma membrane but enclosed by the cell wall. The contents of this special milieu exterior could be regulated by the plasma membrane from the internal, and by the cell wall from the external environment of the cell. Unlike the gram-negative organism, the presence of this space in gram-positive bacteria is still controversial because it cannot be clearly demonstrated. We have shown the importance of some periplasmic bodies in the secretion of penicillinase from Bacillus licheniformis.In negatively stained specimens prepared by a modified technique (Figs. 1 and 2), periplasmic space (PS) contained two kinds of structures: (i) fibrils (F, 100 Å) running perpendicular to the cell wall from the protoplast and (ii) an array of vesicles of various sizes (V), which seem to have evaginated from the protoplast.

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Gram-positive bacteria cell wall-derived lipoteichoic acid induces inflammatory alveolar bone loss through prostaglandin E production in osteoblasts

Scientific Reports ◽

10.1038/s41598-021-92744-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Tsukasa Tominari ◽

Ayumi Sanada ◽

Ryota Ichimaru ◽

Chiho Matsumoto ◽

Michiko Hirata ◽

...

Keyword(s):

Cell Wall ◽

Bone Loss ◽

Alveolar Bone ◽

Osteoclast Differentiation ◽

Lipoteichoic Acid ◽

Alveolar Bone Loss ◽

Gram Negative Bacteria ◽

Gram Positive ◽

Gram Negative ◽

Gram Positive Bacteria

AbstractPeriodontitis is an inflammatory disease associated with severe alveolar bone loss and is dominantly induced by lipopolysaccharide from Gram-negative bacteria; however, the role of Gram-positive bacteria in periodontal bone resorption remains unclear. In this study, we examined the effects of lipoteichoic acid (LTA), a major cell-wall factor of Gram-positive bacteria, on the progression of inflammatory alveolar bone loss in a model of periodontitis. In coculture of mouse primary osteoblasts and bone marrow cells, LTA induced osteoclast differentiation in a dose-dependent manner. LTA enhanced the production of PGE2 accompanying the upregulation of the mRNA expression of mPGES-1, COX-2 and RANKL in osteoblasts. The addition of indomethacin effectively blocked the LTA-induced osteoclast differentiation by suppressing the production of PGE2. Using ex vivo organ cultures of mouse alveolar bone, we found that LTA induced alveolar bone resorption and that this was suppressed by indomethacin. In an experimental model of periodontitis, LTA was locally injected into the mouse lower gingiva, and we clearly detected alveolar bone destruction using 3D-μCT. We herein demonstrate a new concept indicating that Gram-positive bacteria in addition to Gram-negative bacteria are associated with the progression of periodontal bone loss.

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