Abstract:The antimicrobial peptide nisin exerts its activity by a unique dual mechanism. It permeates the cell membranes of Gram-positive bacteria by binding to the cell wall precursor Lipid II and inhibits cell wall synthesis. Binding of nisin to Lipid II induces the formation of large nisin-Lipid II aggregates in the membrane of bacteria as well as in Lipid II-doped model membranes. Mechanistic details of the aggregation process and its impact on membrane permeation are still unresolved. In our experiments, we found … Show more
“…Class 1 lantibiotics are known to bind to lipid II by forming a cage around the pyrophosphate residue using rings A and B (the lipid II binding domain) (47,48). Furthermore, the latter half of the peptide is believed to help in the lateral assembly of the lantibiotic-lipid II complexes to form islands (17,49). In nisin, these islands form a pore complex, but the epidermin group primarily sequesters lipid II without forming a pore, as has been reported in fluorescently labeled lipid II vesicle experiments (17,50).…”
Mutacin 1140 belongs to the epidermin group of lantibiotics. Epidermin class lantibiotics are ribosomally synthesized and posttranslationally modified antibiotics with potent activity against Gram-positive bacteria. In particular, this class is effective at targeting drug-resistant Streptococcus pneumoniae, methicillin-resistant Staphylococcus aureus (MRSA), Mycobacterium tuberculosis, and Clostridium difficile.residue is derived from a decarboxylation of a terminal cysteine that is involved in lanthionine ring formation. Studies on mutacin 1140 have revealed new insight into the structural importance of the C-terminal AviCys residue. A C-terminal carboxyl analogue of mutacin 1140 was engineered. Capping the C-terminal carboxyl group with a primary amine restores bioactivity and affords a novel opportunity to synthesize new analogues. A C-terminal fluorescein-labeled mutacin 1140 analogue traps lipid II into a large lipid II lantibiotic complex, similar to that observed in vivo for the lantibiotic nisin. A C-terminal carboxyl analogue of mutacin 1140 competitively inhibits the activity of native mutacin 1140 and nisin. The presence of a C-terminal carboxyl group prevents the formation of the large lipid II lantibiotic complexes but does not prevent the binding of the lantibiotic to lipid II.
IMPORTANCE This study addressed the importance of the C-terminal S-[(Z)-2-aminovinyl]-D-cysteine (AviCys) residue for antibacterial activity. We have learned that the posttranslational modification for making the AviCys residue is presumably important for the lateral assembly mechanism of activity that traps lipid II into a large complex. The C-terminal carboxyl analogue of this class of lantibiotics is agreeable to the addition of a wide variety of substrates. The addition of fluorescein enabled in vivo visualization of the epidermin class of lantibiotics in action. These results are significant because, as we demonstrate, the presence of the AviCys residue is not essential for bioactivity, but, more importantly, the removal of the carboxyl group is essential. The ability to make a C-terminal carboxyl analogue that is modifiable will facilitate the synthesis of novel analogues of the epidermin class of lantibiotics that can be developed for new applications.KEYWORDS lantibiotic, decarboxylation, mutacin 1140, nisin L antibiotics, or lanthionine-containing antibiotics, are characterized by their posttranslational modifications (PTMs) (1). Dehydrations of serine and threonine residues into dehydroalanine and dehydrobutyrine residues, respectively, are a common modification found in lantibiotics. These dehydrated residues can be cyclized with cysteines to form thioether bridges, which are called lanthionines (2, 3). The unmodified lantibiotic is encoded by the gene lanA. The genes for biosynthesis and regulation are generally grouped together in a biosynthetic gene cluster. In class I lantibiotics, dehydrations are carried out by the dehydratase, LanB. Lanthionine ring formation is catalyzed by LanC. Lantibiotics can ...
“…Class 1 lantibiotics are known to bind to lipid II by forming a cage around the pyrophosphate residue using rings A and B (the lipid II binding domain) (47,48). Furthermore, the latter half of the peptide is believed to help in the lateral assembly of the lantibiotic-lipid II complexes to form islands (17,49). In nisin, these islands form a pore complex, but the epidermin group primarily sequesters lipid II without forming a pore, as has been reported in fluorescently labeled lipid II vesicle experiments (17,50).…”
Mutacin 1140 belongs to the epidermin group of lantibiotics. Epidermin class lantibiotics are ribosomally synthesized and posttranslationally modified antibiotics with potent activity against Gram-positive bacteria. In particular, this class is effective at targeting drug-resistant Streptococcus pneumoniae, methicillin-resistant Staphylococcus aureus (MRSA), Mycobacterium tuberculosis, and Clostridium difficile.residue is derived from a decarboxylation of a terminal cysteine that is involved in lanthionine ring formation. Studies on mutacin 1140 have revealed new insight into the structural importance of the C-terminal AviCys residue. A C-terminal carboxyl analogue of mutacin 1140 was engineered. Capping the C-terminal carboxyl group with a primary amine restores bioactivity and affords a novel opportunity to synthesize new analogues. A C-terminal fluorescein-labeled mutacin 1140 analogue traps lipid II into a large lipid II lantibiotic complex, similar to that observed in vivo for the lantibiotic nisin. A C-terminal carboxyl analogue of mutacin 1140 competitively inhibits the activity of native mutacin 1140 and nisin. The presence of a C-terminal carboxyl group prevents the formation of the large lipid II lantibiotic complexes but does not prevent the binding of the lantibiotic to lipid II.
IMPORTANCE This study addressed the importance of the C-terminal S-[(Z)-2-aminovinyl]-D-cysteine (AviCys) residue for antibacterial activity. We have learned that the posttranslational modification for making the AviCys residue is presumably important for the lateral assembly mechanism of activity that traps lipid II into a large complex. The C-terminal carboxyl analogue of this class of lantibiotics is agreeable to the addition of a wide variety of substrates. The addition of fluorescein enabled in vivo visualization of the epidermin class of lantibiotics in action. These results are significant because, as we demonstrate, the presence of the AviCys residue is not essential for bioactivity, but, more importantly, the removal of the carboxyl group is essential. The ability to make a C-terminal carboxyl analogue that is modifiable will facilitate the synthesis of novel analogues of the epidermin class of lantibiotics that can be developed for new applications.KEYWORDS lantibiotic, decarboxylation, mutacin 1140, nisin L antibiotics, or lanthionine-containing antibiotics, are characterized by their posttranslational modifications (PTMs) (1). Dehydrations of serine and threonine residues into dehydroalanine and dehydrobutyrine residues, respectively, are a common modification found in lantibiotics. These dehydrated residues can be cyclized with cysteines to form thioether bridges, which are called lanthionines (2, 3). The unmodified lantibiotic is encoded by the gene lanA. The genes for biosynthesis and regulation are generally grouped together in a biosynthetic gene cluster. In class I lantibiotics, dehydrations are carried out by the dehydratase, LanB. Lanthionine ring formation is catalyzed by LanC. Lantibiotics can ...
“…[7] As econd effect of this binding is the inhibition of peptidoglycan biosynthesis, caused by the large-scale sequestration and aggregation of lipid II. [8,9] The importance of the interaction with lipid II in the antibacterial action of nisin has also been demonstrated in functional studies, in which it wass hown that nisin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)i sa ble to antagonize the activity of WT nisin. [10] NMR studies have provedt ob eav aluable method in the study of lantibiotic conformation and lipid II binding.…”
Section: Introductionmentioning
confidence: 90%
“…[46][47][48][49] We have previously reported the solid-phase synthesis of the individual Aa nd Br ings of nisin and of the related lantibiotic, mutacin I, [25] and have investigated the conformationalp roperties of these isolated rings ands ynthetic analogues by NMR. We have now extended this work to prepares ynthetic analogues of WT nisin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12), which can be compared with WT nisin(1-12) itself (1)( Figure 2). (Thr2, Ser5) analogue 2 was designed using the amino acids in the 2-and5 -positions that would be present in the biosynthetic precursor peptide, and that would undergo dehydration by the enzyme NisB in the producing organism.…”
Section: Peptides Ynthesismentioning
confidence: 99%
“…[34] As parto ft his work the authors also investigated the effect of replacing both Dha and Dhb with Ala, and found that the presence of the dehydro residues increased the affinity of the dicarbab ridged peptides for lipid II. Introduction of at hirdc yclic constraint in dicarbab ridged analogueso fn isin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12), by creating al actam bridge between the N-terminus and the Br ing, has also been investigated by Harmsen et al [35] The resulting reduction in flexibility increased the affinity of the peptide for lipid II over the bicyclic dicarba analogue, but was still five-fold less active than WT nisin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12).…”
Section: Introductionmentioning
confidence: 99%
“…The first aim of this research was to furtherd evelopo ur existing solid-phase peptides ynthesis (SPPS) methodology to enable the synthesis of two analogues of nisin(1-12) bearing dehydro residue replacementsa nd aM eLan!Lans ubstitution. Our second aim was to examine in detail the solution conformations of these peptides using the NAMFIS method and compare them to the conformations of WT nisin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)i no rder to explore the effect of residue mutation on ring conformation and pre-organization for lipid II binding. Our third aim was to carry out ad etailed conformational analysiso fW Tn isin (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12) itself, in order to elucidatef urtherh ow this peptides equence achievesh ighly selectiveb inding to lipid II, and to compare the conformations to the previously published structures of full length nisin, both alone and bound to lipid II.…”
Natural products that target lipid II, such as the lantibiotic nisin, are strategically important in the development of new antibacterial agents to combat the rise of antimicrobial resistance. Understanding the structural factors that govern the highly selective molecular recognition of lipid II by the N‐terminal region of nisin, nisin(1–12), is a crucial step in exploiting the potential of such compounds. In order to elucidate the relationships between amino acid sequence and conformation of this bicyclic peptide fragment, we have used solid‐phase peptide synthesis to prepare two novel analogues of nisin(1–12) in which the dehydro residues have been replaced. We have carried out an NMR ensemble analysis of one of these analogues and of the wild‐type nisin(1–12) peptide in order to compare the conformations of these two bicyclic peptides. Our analysis has shown the effects of residue mutation on ring conformation. We have also demonstrated that the individual rings of nisin(1–12) are pre‐organised to an extent for binding to the pyrophosphate group of lipid II, with a high degree of flexibility exhibited in the central amide bond joining the two rings.
Chalcones, valuable precursors for flavonoids, have important antibacterial and antifungal activities against bacteria, pathogens, harmful fungi and even antibiotic-resistant microorganisms that cause food spoilage and infectious diseases. It is widely known that chalcones target various vital metabolic pathways of the bacterial cells, but little is known about their action on the cell membrane integrity. In the present study, we studied the antibacterial activity of 12 different substituted chalcones in a comparative way and revealed that the phenolic chalcones are superior to other substituted derivatives against both Gram-negative and Gram-positive bacteria. We also demonstrate that the cell membrane is the first barrier that the chalcone molecules face for their action, and that phenolic chalcones increase ionic cell membrane permeability to a greater extent than the other substituted members. Especially, ion leakage can be detected at lower concentrations than the minimum inhibitory levels against Gram-positive bacteria. Phenolic chalcones are superior to other substituted derivatives in their antibacterial action and cause leakage of ions from Gram-positive bacteria even in concentrations lower than the inhibitory levels. Ion leakage from Gram-positive bacterial cytoplasm is prior to the membrane deformation and cell death. Thus, we propose that ion leakage contribute to the greater activity of phenolic chalcones in comparison to non-phenolic ones, on Gram-positive bacteria. Even though, disruption of metabolic pathways may be the principal mode of action of chalcones; in accord with our observations, we propose that the ion leakage precedes other inhibitory effects and contribute to the antibacterial action of phenolic chalcones.
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