Dissection and Modulation of the Four Distinct Activities of Nisin by Mutagenesis of Rings A and B and by C-Terminal Truncation

• Published in: Applied and Environmental Microbiology Vol. 73; no. 18; pp. 5809 - 5816
• Main Authors: Rink, Rick, Wierenga, Jenny, Kuipers, Anneke, Kluskens, Leon D, Driessen, Arnold J.M, Kuipers, Oscar P, Moll, Gert N
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01-09-2007; American Society for Microbiology (ASM)
• Subjects: Amino acids; Anti-Bacterial Agents - biosynthesis; Anti-Bacterial Agents - chemistry; Antibiotics; Bacillus; Bacteria; Bacteria - drug effects; Bacteria - growth & development; Bacterial Proteins; Biological and medical sciences; Enzymology and Protein Engineering; Fundamental and applied biological sciences. Psychology; Immune system; Lactococcus lactis; Microbiology; Mutagenesis; Mutagenesis, Site-Directed; Nisin - biosynthesis; Nisin - chemistry; Nisin - genetics; Nisin - pharmacology; Peptides; Protein Conformation; Protein Engineering - methods; Recombinant Proteins - chemistry; Recombinant Proteins - pharmacology; Structure-Activity Relationship; Nisin; Mutagenesis; Modulation; Bacteriocin
• ISSN: 0099-2240; 1098-5336
• DOI: 10.1128/AEM.01104-07

Nisin A is a pentacyclic antibiotic peptide produced by various Lactococcus lactis strains. Nisin displays four different activities: (i) it autoinduces its own synthesis; (ii) it inhibits the growth of target bacteria by membrane pore formation; (iii) it inhibits bacterial growth by interfering with cell wall synthesis; and, in addition, (iv) it inhibits the outgrowth of spores. Here we investigate the structural requirements and relevance of the N-terminal thioether rings of nisin by randomization of the ring A and B positions. The data demonstrate that: (i) mutation of ring A results in variants with enhanced activity and a modulated spectrum of target cells; (ii) for the cell growth-inhibiting activity of nisin, ring A is rather promiscuous with respect to its amino acid composition, whereas the bulky amino acid residues in ring B abolish antimicrobial activity; (iii) C-terminally truncated nisin A mutants lacking rings D and E retain significant antimicrobial activity but are unable to permeabilize the target membrane; (iv) the dehydroalanine in ring A is not essential for the inhibition of the outgrowth of Bacillus cells; (v) some ring A mutants have significant antimicrobial activities but have decreased autoinducing activities; (vi) the opening of ring B eliminates antimicrobial activity while retaining autoinducing activity; and (vii) some ring A mutants escape the nisin immune system(s) and are toxic to the nisin-producing strain NZ9700. These data demonstrate that the various activities of nisin can be engineered independently and provide a basis for the design and synthesis of tailor-made analogs with desired activities.