Elucidation of the Antimicrobial Mechanism of Mutacin 1140

Smith, Linda L.; Hasper, Hester E.; Breukink, Eefjan; Novák, Jan; Cerkasov, Jirri; Hillman, Jeffrey D.; Wilson-Stanford, Shawanda; Orugunty, Ravi S.

doi:10.1021/bi701262z

Cited by 65 publications

(55 citation statements)

References 33 publications

(58 reference statements)

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“…The lantibiotic nisin shares structural homology with the lanthionine rings A and B. The latter half of the epidermin and nisin peptide is referred to as the lateral assembly domain, which presumably abducts lipid II into large lipid II/lantibiotic complexes (17)(18)(19)(20).…”

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Carboxyl Analogue of Mutacin 1140, a Scaffold for Lead Antibacterial Discovery

Escano

Ravichandran

Salamat

et al. 2017

Appl Environ Microbiol

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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 ...

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Carboxyl Analogue of Mutacin 1140, a Scaffold for Lead Antibacterial Discovery

Escano

Ravichandran

Salamat

et al. 2017

Appl Environ Microbiol

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“…Mutacin 1140 and nisin have structurally similar rings A and B (Fig. 1A), and both nisin and mutacin 1140 bind and abduct lipid II (8)(9)(10)(11)(12)(13). Mutacin 1140 has also been shown to be bactericidal to pathogenic bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), with no significant development of resistance (14).…”

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“…However, the core peptide of mutacin 1140 is 12 amino acids shorter than that of nisin. Mutacin 1140 has the same ability to bind to lipid II, but it does not form bacterial membrane pores like nisin does (12,13).…”

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Biosynthesis and Transport of the Lantibiotic Mutacin 1140 Produced by Streptococcus mutans

et al. 2015

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Lantibiotics are ribosomally synthesized peptide antibiotics composed of an N-terminal leader peptide that is cleaved to yield the active antibacterial peptide. Significant advancements in molecular tools that promote the study of lantibiotic biosynthesis can be used in Streptococcus mutans. Herein, we further our understanding of leader peptide sequence and core peptide structural requirements for the biosynthesis and transport of the lantibiotic mutacin 1140. Our study on mutacin 1140 biosynthesis shows a dedicated secondary cleavage site within the leader peptide and the dependency of transport on core peptide posttranslational modifications (PTMs). The secondary cleavage site on the leader peptide is found at the ؊9 position, and secondary cleavage occurs before the core peptide is transported out of the cell. The coordinated cleavage at the ؊9 position was absent in a lanT deletion strain, suggesting that the core peptide interaction with the LanT transporter enables uniform cleavage at the ؊9 position. Following transport, the LanP protease was found to be tolerant to a wide variety of amino acid substitutions at the primary leader peptide cleavage site, with the exception of arginine at the ؊1 position. Several leader and core peptide mutations produced core peptide variants that had intermediate stages of PTM enzyme modifications, supporting the concept that PTM enzyme modifications, secondary cleavage, and transport are occurring in a highly coordinated fashion. IMPORTANCEMutacin 1140 belongs to the class I lantibiotic family of ribosomally synthesized and posttranslationally modified peptides (RiPPs). The biosynthesis of mutacin 1140 is a highly efficient process which does not lead to a discernible level of production of partially modified core peptide variants. The products isolated from an extensive mutagenesis study on the leader and core peptides of mutacin 1140 show that the posttranslational modifications (PTMs) on the core peptide occur under a highly coordinated dynamic process. PTMs are dictated by the distance of the core peptide modifiable residues from PTM enzyme active sites. The formation of lanthionine rings aids in the formation of successive PTMs, as was observed in a peptide variant lacking a Cterminal decarboxylation. Lantibiotics are a class of ribosomally synthesized peptide antibiotics produced by Gram-positive bacteria, such as Lactococcus lactis and Streptococcus mutans (1, 2). Lantibiotics are characterized by the presence of posttranslational modifications (PTMs), such as dehydrated residues and lanthionine rings. The modified residues 2,3-didehydroalanine (Dha) and 2,3-didehydrobutyrine (Dhb) are formed from dehydration of serines and threonines, respectively. The cyclization between a cysteine and either a Dha or a Dhb forms a lanthionine or a methyllanthionine ring, respectively. The biosynthetic gene cluster contains all the genes necessary to produce a lantibiotic. The lan operon for class I lantibiotics contains genes encoding the lantibiotic peptide (lanA); the...

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“…Nisin, one of the most studied lanthipeptides, is produced by Lactococcus lactis and has been used in the food industry for over 50 years. It was discovered that both nisin and mutacin 1140 abduct lipid II from the site of new cell wall synthesis, ultimately causing cell death (10)(11)(12)(13). Rings A and B provide the structural motif for binding to lipid II and are referred to as the lipid II binding domain (14).…”

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Site-Directed Mutations in the Lanthipeptide Mutacin 1140

Chen

Wilson-Stanford

Cromwell

et al. 2013

Appl Environ Microbiol

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The oral bacterium Streptococcus mutans, strain JH1140, produces the antibiotic mutacin 1140. Mutacin 1140 belongs to a group of antibiotics called lanthipeptides. More specifically, mutacin 1140 is related to the epidermin type A(I) lanthipeptides. Mutagenesis experiments of this group of lanthipeptides have been primarily restricted to the posttranslationally modified mesolanthionine and 3-methyllanthionine residues. Site-directed mutagenesis of the core peptide of mutacin 1140 was performed using the suicide vector pVA891. Substitutions of the N-terminal residue, the charged residue in the hinge region, and residues in ring A and intertwined rings C and D were investigated. A truncation and insertion of residues in ring A and intertwined rings C and D were also performed to determine whether or not they would alter the antimicrobial activity of the producing strain. Bioassays revealed that five of 14 mutants studied had improved antimicrobial activity against the indicator strain Micrococcus luteus ATCC 10240. MICs against Streptococcus mutans UA159, Streptococcus pneumoniae ATCC 27336, Staphylococcus aureus ATCC 25923, Clostridium difficile UK1, and Micrococcus luteus ATCC 10240 were determined for three mutacin 1140 variants that had the most significant increases in bioactivity in the M. luteus bioassay. This mutagenesis study of the epidermin group of lanthipeptides shows that antimicrobial activity can be significantly improved.

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Elucidation of the Antimicrobial Mechanism of Mutacin 1140

Cited by 65 publications

References 33 publications

Carboxyl Analogue of Mutacin 1140, a Scaffold for Lead Antibacterial Discovery

Carboxyl Analogue of Mutacin 1140, a Scaffold for Lead Antibacterial Discovery

Biosynthesis and Transport of the Lantibiotic Mutacin 1140 Produced by Streptococcus mutans

Site-Directed Mutations in the Lanthipeptide Mutacin 1140

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