Frank J. M. Van De Ven

NMR and circular dichroism studies of the lantibiotic nisin in non-aqueous environments

The lantibiotic, nisin, which is known to interact with membranes of certain Gram‐positive bacteria, was studied in three model systems which mimic a membrane‐like environment, i.e. a mixture of trifluoroethanol and water, or micelles of sodium dodecyl sulfate or dodecylphosphocholine. The 1H NMR spectra of nisin in the non‐aqueous environments, at 40°C and pH 3.5, have been assigned completely. The CD and NMR results indicate that the conformation of nisin in the three non‐aqueous environments differs from that in aqueous solution, and that the conformation in the two micellar systems is similar. The major conformational changes, relative to nisin in aqueous solution, occur in the N‐terminus.

Local helix content in an alanine-rich peptide as determined by the complete set of 3JHNα coupling constants

SummaryAlanine-rich peptides serve as models for exploring the factors that control helix structure in peptides and proteins. Scalar CαH-NH couplings (3JHNα) are an extremely useful measure of local helix content; however, the large alanine content in these peptides leads to significant signal overlap in the CαH region of 1H 2D NMR spectra. Quantitative determination of all possible 3JHNα values is, therefore, very challenging. Szyperski and co-workers [(1992) J. Magn. Reson., 99, 552–560] have recently developed a method for determining 3JHNα from NOESY spectra. Because 3JHNα may be determined from 2D peaks outside of the CαH region, there is a much greater likelihood of identifying resolved resonances and measuring the associated coupling constants. It is demonstrated here that 3JHNα can be obtained for every residue in the helical peptide Ac-(AAAAK)3A-NH2. The resulting 3JHNα profile clearly identifies a helical structure in the middle of the peptide and further suggests that the respective helix termini unfold via distinct pathways.

NMR Studies of Lantibiotics: The Three-Dimensional Structure of Nisin in Aqueous Solution

Nisin, a bacteriocin produced by Lactococcus lactis ssp., is a post-translationally modified pentacyclic polypeptide of 34 amino acids. It is a member of the class of bacteriocins, known as lantibiotics, that contain the unusual amino acid lanthionine. Its structure in aqueous solution has been determined on the basis of data obtained from Nuclear Magnetic Resonance Spectroscopy (NMR) studies. Translation of the interproton distance constraints, derived from Nuclear Overhauser Enhancement Spectroscopy (NOESY)l, and torsion angle constraints, derived from Double Quantum Filtered Correlated Spectroscopy (DQF-COSY), into a 3D structure was carried out with the distance geometry program DISMAN, followed by restrained energy minimization using CHARMm. Due to the internal mobility of the polypeptide chain a determination of the precise overal folding of the molecule was prohibited, but parts of the structure could be obtained albeit with sometimes low resolution. The structure of nisin can best be described as follows: the outermost N-and C-terminal regions appear quite flexible, the remainder of the molecule consists of an amphiphilic N-terminal fragment (residues 3–19), joined by a flexible ‘hinge’ region to a rigid doublering fragment formed by residues 23–28. The latter fragment has the appearance of a somewhat overwound α-helix. It is postulated that i) the coupling between residues 23 and 26 as well as between 25 and 28 by thioether bridges and ii) the inversion of the Cα chiralities at positions 23 and 25 occurs via an intermediate α-helical structure of the prenisin molecule