STRUCTURE-FUNCTION RELATIONS OF VARIANT AND FRAGMENT NISINS STUDIED WITH MODEL MEMBRANE SYSTEMS

Nisin, a 34 residue lantibiotic produced by strains of Lactococcus lac tis subsp. lactis, exerts antimicrobial activity against Gram-positive bacteria at the cytoplasmic membrane. The structural aspects of nisin which facilitate membrane interaction and permeabilization have been investigated in planar lipid bilayers and liposomes with proteolytic f ragments and site-directed variants. N-Terminal nisin fragments N1-12 and N1-20 had little effect on phospholipid mobility, on macroscopic e lectrical conductance, or on calcein release from liposomes. By contra st, the I30W nisin A variant induced a time-dependent reduction in lip id mobility, indicative of nisin-membrane surface interactions, as wel l as a decline in membrane capacitance, rise in conductance, and calce in release from liposomes. In these respects I30W nisin A is similar t o native nisin. Charge substitutions were also engineered to generate K12L and H27K nisin A variants, both of which were similar to I30W nis in A with respect to an overall reduction in phospholipid mobility. Wh ile the K12L nisin A variant elicited a higher increase in membrane ca pacitance and electrical conductance than I30W nisin A, the H27K nisin A variant elicited weaker effects. These results point to a substanti al role for intramembrane charged residues in controlling ion flow thr ough nisin-doped membranes. Native nisin and variants elicit an enhanc ed release of calcein from liposomes composed of the negatively-charge d phospholipids cardiolipin and phosphatidylserine, compared with phos pholipid bearing no net charge, suggesting that an electrostatic attra ction encourages the initial nisin-membrane association. The results a re discussed in the context of other recently proposed models for nisi n action.