The role of lipid II in membrane binding of and pore formation by nisin analyzed by two combined biosensor techniques.
Nisin, a peptide antibiotic, efficiently kills bacteria through a unique mechanism which includes inhibition of cell wall biosynthesis and pore formation in cytoplasmic membranes. Both mechanisms are based on interaction with the cell wall precursor lipid II which is simultaneously used as target and pore constituent. We combined two biosensor techniques to investigate the nisin activity with respect to membrane binding and pore formation in real time. Quartz crystal microbalance (QCM) allows the detection of nisin binding kinetics. The presence of 0.1 mol% lipid II strongly increased nisin binding affinity to DOPC (k(D) 2.68 x 10(-7) M vs. 1.03 x 10(-6) M) by a higher association rate. Differences were less pronounced while using negatively charged DOPG membranes. However, lipid II does not influence the absolute amount of bound nisin. Cyclic voltammetry (CV) data confirmed that in presence of 0.1 mol% lipid II, nanomolar nisin concentrations were sufficient to form pores, while micromolar concentrations were necessary in absence of lipid II. Both techniques suggested unspecific destruction of pure DOPG membranes by micromolar nisin concentrations which were prevented by lipid II. This model membrane stabilization by lipid II was confirmed by atomic force microscopy. Combined CV and QCM are valuable to interpret the role of lipid II in nisin activity.
Christ
Katrin
K
University of Bonn, Department of Pharmacy, An der Immenburg 4, 53121 Bonn, Germany. kchrist@uni-bonn.de
Wiedemann
Imke
I
Bakowsky
Udo
U
Sahl
Hans-Georg
HG
Bendas
Gerd
G
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D015374:Biosensing Techniques / Q000379:methods*; D036681:Fluorescence Recovery After Photobleaching; D007700:Kinetics; D008566:Membranes / Q000737:chemistry / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009561:Nisin / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010713:Phosphatidylcholines / Q000378:metabolism; D010715:Phosphatidylglycerols / Q000378:metabolism; D014538:Uridine Diphosphate N-Acetylmuramic Acid / Q000031:analogs & derivatives* / Q000302:isolation & purification / Q000378:metabolism
D000900:Anti-Bacterial Agents; D010713:Phosphatidylcholines; D010715:Phosphatidylglycerols; D014538:Uridine Diphosphate N-Acetylmuramic Acid; C069249:muramyl-NAc-(pentapeptide)pyrophosphoryl-undecaprenol; D009561:Nisin; C051388:1,2-dioleoyl-sn-glycero-3-phosphoglycerol; C017251:1,2-oleoylphosphatidylcholine