Structure-function relations of variant and fragment nisins studied with model membrane systems.
Nisin, a 34 residue lantibiotic produced by strains of Lactococcus lactis 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 fragments and site-directed variants. N-Terminal nisin fragments N1-12 and N1-20 had little effect on phospholipid mobility, on macroscopic electrical conductance, or on calcein release from liposomes. By contrast, the I30W nisin A variant induced a time-dependent reduction in lipid mobility, indicative of nisin-membrane surface interactions, as well as a decline in membrane capacitance, rise in conductance, and calcein release from liposomes. In these respects I30W nisin A is similar to native nisin. Charge substitutions were also engineered to generate K12L and H27K nisin A variants, both of which were similar to I30W nisin A with respect to an overall reduction in phospholipid mobility. While the K12L nisin A variant elicited a higher increase in membrane capacitance and electrical conductance than I30W nisin A, the H27K nisin A variant elicited weaker effects. These results point to a substantial role for intramembrane charged residues in controlling ion flow through nisin-doped membranes. Native nisin and variants elicit an enhanced release of calcein from liposomes composed of the negatively-charged phospholipids cardiolipin and phosphatidylserine, compared with phospholipid bearing no net charge, suggesting that an electrostatic attraction encourages the initial nisin-membrane association. The results are discussed in the context of other recently proposed models for nisin action.
Giffard
C J
CJ
Department of Biology, University of York, U.K. c.giffard@uea.ac.uk
Dodd
H M
HM
Horn
N
N
Ladha
S
S
Mackie
A R
AR
Parr
A
A
Gasson
M J
MJ
Sanders
D
D
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism; D002308:Cardiolipins / Q000378:metabolism; D005452:Fluoresceins / Q000378:metabolism; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000378:metabolism; D007700:Kinetics; D013294:Lactococcus lactis / Q000737:chemistry*; D008051:Lipid Bilayers / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D009561:Nisin / Q000737:chemistry* / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D010539:Permeability; D010714:Phosphatidylethanolamines / Q000378:metabolism; D013329:Structure-Activity Relationship
D000900:Anti-Bacterial Agents; D002308:Cardiolipins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008081:Liposomes; D010446:Peptide Fragments; D010714:Phosphatidylethanolamines; D009561:Nisin; C042077:N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)phosphatidylethanolamine; C007740:fluorexon