Solution structure of microcin J25, the single macrocyclic antimicrobial peptide from Escherichia coli

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Abstract
The three-dimensional solution structure of microcin J25, the single cyclic representative of the microcin antimicrobial peptide class produced by enteric bacteria, was determined using two-dimensional 1H NMR spectroscopy and molecular modeling. This hydrophobic 21-residue peptide exhibits potent activity directed to Gram-negative bacteria. Its primary structure, cyclo(-V1GIGTPISFY10GGGAGHVPEY20F-), has been determined previously [Blond, A., Péduzzi, J., Goulard, C., Chiuchiolo, M. J., Barthélémy, M., Prigent, Y., Salomón, R.A., Farías, R.N., Moreno, F. & Rebuffat, S. (1999) Eur. J. Biochem., 259, 747–755]. Conformational parameters (3JNHCαH coupling constants, quantitative nuclear Overhauser enhancement data, chemical shift deviations, temperature coefficients of amide protons, NH–ND exchange rates) were obtained in methanol solution. Structural restraints consisting of 190 interproton distances inferred from NOE data, 11 φ backbone dihedral angle and 9 χ1 angle restraints derived from the coupling constants and three hydrogen bonds in agreement with the amide exchange rates were used as input for simulated annealing calculations and energy minimization in the program x plor. Microcin J25 adopts a well-defined compact structure consisting of a distorted antiparallel β sheet, which is twisted and folded back on itself, thus resulting in three loops. Residues 7–10 and 17–20 form the more regular part of the β sheet. The region encompassing residues Gly11–His16 consists of a distorted β hairpin, which divides into two small loops and is stabilized by an inverse γ turn and a type I′β turn. The reversal of the chain leading to the Phe21–Pro6 loop results from a mixed β/γ turn. A cavity, in which the hydrophilic Ser8 side-chain is confined, is delimited by two crab pincer-like regions that comprise residues 6–8 and 18–1.