The key to Nisin's antibacterial activity

Nisin, an antimicrobial peptide produced by lactic acid bacteria, owes its potent bacteriostatic activity—particularly against Gram-positive bacteria—to its unique molecular structure. The formation of thioether bonds and the stability of internal cyclic structures are core elements determining its function. These structural features not only endow Nisin with a specific spatial conformation but also directly participate in the mechanism of action against target microorganisms.

I. Thioether Bonds: The "Backbone" of Nisin’s Molecular Structure

Nisin’s amino acid sequence contains 5 rare lanthionine-type residues (e.g., lanthionine, β-methyl lanthionine), which are linked via thioether bonds (-S-) between cysteine (Cys) and dehydroalanine (Dha) or dehydrobutyrine (Dhb). In Nisin A (the most common subtype), 34 amino acid residues form 5 distinct cyclic systems through 5 thioether bonds: Ring A (residues 8–11), Ring B (12–19), Ring C (20–26), Ring D (27–31), and Ring E (32–34).

The formation of thioether bonds is a key post-translational modification step in Nisin biosynthesis: specific enzymes catalyze the addition reaction between the thiol group (-SH) of cysteine and the double bond of dehydroamino acids, folding the linear peptide chain into a rigid cyclic structure. This covalent bonding grants Nisin exceptional stability—it resists degradation by proteases (e.g., pepsin, trypsin) and maintains conformational integrity across a wide pH range (2–6) and at high temperatures (e.g., pasteurization temperatures), making it highly advantageous for applications such as food preservation.

II. Internal Cyclic Structures: The Foundation for Spatial Arrangement of Functional Sites

The internal cyclic structures formed by thioether bonds are not merely structural supports; their precise spatial arrangement exposes key functional sites, directly participating in interactions with target bacteria. Nisin’s antimicrobial mechanism primarily relies on binding to lipid II (a peptidoglycan precursor) on bacterial cell membranes, inhibiting cell wall synthesis, and forming pores to cause intracellular substance leakage. The spatial conformation of the cyclic structures is critical to this process.

Synergistic role of Rings B and C: Studies indicate that Ring B (residues 12–19) and Ring C (20–26) are core regions for lipid II binding. The lysine residue (Lys22) in Ring C binds to the pyrophosphate group of lipid II via electrostatic interactions, while the rigid structure of Ring B provides spatial positioning, ensuring the binding site matches lipid II spatially. If thioether bond cleavage disrupts Rings B or C, Nisin’s lipid II binding capacity decreases by over 80%, and antimicrobial activity is nearly lost.

Membrane perforation function of Ring E: Though small, Ring E (residues 32–34) is crucial for pore formation. Its compact structure (only 3 amino acids) inserts into the phospholipid bilayer of bacterial cell membranes, collaborating with other rings to induce membrane disorder. If Ring E’s thioether bond is disrupted (e.g., by chemical modification blocking cysteine thiols), Nisin can still bind lipid II but fails to form effective pores, reducing bacteriostatic activity to 10%–20% of its original level.

Balanced flexibility between rings: The internal rings are not completely rigid; the bond length and angle of thioether bonds grant inter-ring flexibility, allowing Nisin to adapt to curvature differences in cell membranes of various bacteria. For example, the linker region between Rings A and B (residues 11–12) has limited rotatability, adjusting binding posture based on lipid II distribution to enhance broad-spectrum activity against different strains.

III. Correlation Between Structural Integrity and Antimicrobial Activity

Cleavage of thioether bonds or disruption of internal rings directly abolishes Nisin’s antimicrobial activity. For instance, treating Nisin with reducing agents (e.g., dithiothreitol) can reduce some thioether bonds (particularly affecting Rings A and B), linearizing the molecule. This increases the minimum inhibitory concentration (MIC) against Staphylococcus aureus from 0.1 μg/mL to over 50 μg/mL. Genetic engineering replacement of key cysteine residues (e.g., mutating Cys19 to alanine) prevents Ring B formation, rendering Nisin nearly ineffective against Bacillus species.

Moreover, the stability of internal rings affects Nisin’s efficiency. Compared to linear antimicrobial peptides, cyclic Nisin (with thioether bonds) has a higher binding constant (Ka ≈ 10⁸ M⁻¹) for lipid II and dissociates more slowly, continuously blocking cell wall synthesis and disrupting membranes. This efficiency allows it to function at extremely low concentrations (typically 0.01–1 μg/mL), far lower than other natural antimicrobial peptides.

IV. Summary and Application Insights

Thioether bonds, by constructing stable internal rings, endow Nisin with resistance to degradation, target-binding specificity, and membrane-perforating ability—collectively forming the core of its antimicrobial activity. This structure-function relationship provides clear directions for Nisin modification and application: for example, optimizing thioether bond positions via directed mutagenesis to enhance binding to lipid II in specific pathogens, or chemically modifying Ring E to improve hydrophobicity for stronger membrane perforation. Additionally, protecting thioether bonds and ring structures (e.g., controlling pH and temperature during processing) is key to maintaining its efficacy.

A deep understanding of this relationship not only clarifies Nisin’s antimicrobial mechanism but also provides a molecular model for designing new high-efficiency antimicrobial peptides (e.g., artificial peptides mimicking Nisin’s cyclic structure), holding significant value in addressing drug-resistant bacterial infections and developing natural preservatives.