Solution structures of nisin A and its two major degradation products determined by n.m.r

Lian, Lu‐Yun; Chan, Weng C.; Morley, Sam; Roberts, Gordon C. K.; Bycroft, Barrie W.; Jackson, David E.

doi:10.1042/bj2830413

Cited by 77 publications

(53 citation statements)

References 12 publications

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“…We have been studying the structure-activity relationships in nisin and subtilin, by both genetic and chemical modification of the structure [12][13][14][15][16][17][18][19][20], with a view to understanding their mechanism of action and to developing new derivatives with desirable properties. We now report the preparation, characterisation and anti-bacterial activity of a number of proteolytic fragments of nisin and subtilin, which allow us to define the parts of the molecule most important for biological activity.…”

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confidence: 99%

Structure‐activity relationships in the peptide antibiotic nisin: antibacterial activity of fragments of nisin

Chan¹,

Leyland²,

Clark³

et al. 1996

FEBS Letters

123

160

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The post-translationally modified peptide antibiotic nisin has been cleaved by a number of proteases and the fragments produced purified, characterised chemically, and assayed for activity in inhibiting the growth of Lactococcus lactis MG1614 and Micrococcus luteus NCDO8166. These results provide information on the importance of different parts of the nisin molecule for its growth-inhibition activity. Removal of the C-terminal five residues leads to approximately a 10-fold decrease in potency, while removal of a further nine residues, encompassing two of the lanthionine rings, leads to a 100-fold decrease. There are some differences between analogous fragments of nisin and subtilin, suggesting possible subtle differences in mode of action. Cleavage within, or removal of, lanthionine ring C essentially abolishes the activity of nisin. The fragment ulsin t-12 is inactive itself, and specifically antagonises the growth-inhibitory action of nisin. These results are discussed in lerms of current models for the mechanism of action of nisin.Key words: Nisin; Lantibiotic; Peptide antibiotic; I'roteolysis; Structure-activity relationships mation of voltage-dependent pores in biological membranes [8][9][10], although recent evidence indicates that the inhibition of the outgrowth of spores by nisin and subtilin takes place by a different mechanism [11,12]. We have been studying the structure-activity relationships in nisin and subtilin, by both genetic and chemical modification of the structure [12][13][14][15][16][17][18][19][20], with a view to understanding their mechanism of action and to developing new derivatives with desirable properties. We now report the preparation, characterisation and anti-bacterial activity of a number of proteolytic fragments of nisin and subtilin, which allow us to define the parts of the molecule most important for biological activity. Materials and methodsNisin and subtilin were prepared as previously described [14,15]; [Ser33]-nisin, in which the serine residue at position 33 has escaped processing to a dehydroalanine residue, was isolated as a minor component of commercial nisin. All proteases were obtained from Sigma Chemical Co., Poole, Dorset, UK. ~. IntroductionThe post-translationally modified peptide antibiotics known :ts 'lantibiotics' contain cyclic structures formed by lanthioaine or 3-methyl-lanthionine residues, and often also dehydroalanine and/or dehydrobutyrine residues [1]. The first lan-!ibiotic to be characterised was nisin, produced by strains of Lactococcus lactis carrying a transposon containing genes coding for the nisin precursor and for proteins involved in aisin biosynthesis and resistance [2][3][4][5]. Nisin has been quite videly used as a food preservative, notably in cheese and ,~ther dairy products and in canned vegetables, for some 30 .,ears [6,7]. It inhibits the growth of a wide range of Gram~ositive organisms, and also inhibits the germination and/or mtgrowth of spores of Bacillus and Clostridium species [6]. Fhe growth-inhibitory activity of nisi...

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confidence: 99%

Structure‐activity relationships in the peptide antibiotic nisin: antibacterial activity of fragments of nisin

Chan¹,

Leyland²,

Clark³

et al. 1996

FEBS Letters

123

160

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“…The threedimensional structure of nisin appeared to be ill defined from initial NMR data, suggesting adoption of a variable conformation in aqueous solutions. The involvement of two relatively rigid domains (amino acids 3-19 and 23-28), connected by a flexible hinge region, has since been deduced (van de Ven et al, 1991;Lian et al, 1992). Nisin interaction with the membranelike environments of SDS or DodPCho (dodecylphosphocholine) micelles induced a structural transition in the N-terminal domain (first ring) without effect on the overall fold of the molecule (van den Hooven et al, 1993).…”

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Interaction of the Lantibiotic Nisin with Membranes Revealed by Fluorescence Quenching of an Introduced Tryptophan

Martin

Ruysschaert

Sanders

et al. 1996

European Journal of Biochemistry

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Nisin is a lantibiotic produced by strains of Lactococcus lactis subsp. lactis. The target for nisin action is the cytoplasmic membrane of gram-positive bacteria. To aid understanding of its mode of action, the interaction of nisin with vesicles of differing phospholipid composition were investigated by fluorescence techniques, using a variant of nisin in which the isoleucine at position 30 was replaced by a tryptophan residue. Activity of the site-directed variant containing tryptophan was established to be similar to that of the wild-type peptide. Fluorescence experiments showed a blue shift of the emission wavelength maximum in the presence of lipid vesicles, indicating that the tryptophan residue enters a more liydrophobic environment. Quenching experiments with aqueous and membrane-restricted quenchers (iodide and spin-labelled lipids, respectively) both confirmed a non-aqueous environment for the Trp30 residue, and implied that the residue resides between 0.36 nm and 0.52 nm from the centre of the membrane, depending on the lipid identity. The results clearly demonstrate that nisin interacts strongly with the hydrophobic phase of lipid vesicles. This interaction is stronger in the presence of negatively charged lipids suggesting their importance i n the functional interaction of nisin with membranes.

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“…2). Two chemically modified nisin A species, nisin-( 1 -32)-peptide amide and [Ile4-amide,pyruvyl-Leu6Jdes-DhaS-nisin-( 1 -32)-peptide amide, which were formed according to this mechanism, were isolated and characterized by twodimensional NMR techniques (Chan et al, 1989;Lian et al, 1992). It appeared that the removal of the two C-terminal residues does not affect the biological activity.…”

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Structure and Biological Activity of Chemically Modified Nisin A Species

Rollema¹,

Metzger

Both³

et al. 1996

European Journal of Biochemistry

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(Received 21 May112 August 19Y6) -EJB 96 074213Nisin, a 34-residue peptide bacteriocin, contains the less common amino acids lanthionine, p-methyllanthionine, dehydroalanine (Dha), and dehydrobutyrine (Dhb). Several chemically modified nisin A species were purified by reverse-phase HPLC and characterized by two-dimensional NMR and electrospray mass spectrometry. Five constituents, [2-hydroxy-AlaS]nisin, [Tle4-amide,pyruvyl-Leu6Jdes-Dha5-nisin, [Met(0)2l]nisin, [Ser33]nisin, and nisin-(I -32)-peptide amide, were found in a commercial nisin sample. A further species, [2-hydroxy-AIaS]nisin-( 1 -32)-peptide amide, was obtained by freeze drying an acidic nisin solution. These compounds are formed by chemical modification of nisin: the addition of a water molecule to the dehydroalanine residues, which can lead to the cleavage of the polypeptide chain, or the oxidation of methionine residues.The 2-hydroxyalanine-containing products have a limited stability; they are spontaneously converted into the corresponding des-dehydroalanine derivatives. The growth-inhibiting activity of the modified nisins towards different bacteria was determined. The 2-hydroxyalanine-containing species and the desdehydroalanine derivative show a strong reduction in biological activity as compared to native nisin.[Met(0)2l]nisin and [Ser33]nisin show moderate or no reduction in biological activity.

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Solution structures of nisin A and its two major degradation products determined by n.m.r

Cited by 77 publications

References 12 publications

Structure‐activity relationships in the peptide antibiotic nisin: antibacterial activity of fragments of nisin

Structure‐activity relationships in the peptide antibiotic nisin: antibacterial activity of fragments of nisin

Interaction of the Lantibiotic Nisin with Membranes Revealed by Fluorescence Quenching of an Introduced Tryptophan

Structure and Biological Activity of Chemically Modified Nisin A Species

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