Complex formation of amphotericin B in sterol-containing membranes as evidenced by surface plasmon resonance.
Amphotericin B (AmB) is a membrane-active antibiotic that increases the permeability of fungal membranes. Thus, the dynamic process of its interaction with membranes poses intriguing questions, which prompted us to elaborate a quick and reliable method for real-time observation of the drug's binding to phospholipid liposomes. We focused on surface plasmon resonance (SPR) and devised a new modification method of sensor chips, which led to a significant reduction in the level of nonspecific binding of the drug in a control lane. With this method in hand, we examined the affinity of AmB for various membrane preparations. As expected, AmB exhibited much higher affinity for sterol-containing palmitoyloleoylphosphatidylcholine membranes than those without sterol. The sensorgrams recorded under various conditions partly fitted theoretical curves, which were based on three interaction models. Among those, a two-state reaction model reproduced well the sensorgram of AmB binding to an ergosterol-containing membrane; in this model, two states of membrane-bound complexes, AB and AB*, are assumed, which correspond to a simple binding to the surface of the membrane (AB) and formation of another assembly in the membrane (AB*) such as an ion channel complex. Kinetic analysis demonstrated that the association constant in ergosterol-containing POPC liposomes is larger by 1 order of magnitude than that in the cholesterol-containing counterpart. These findings support the previous notion that ergosterol stabilizes the membrane-bound assembly of AmB.
Mouri
Ryota
R
Department of Chemistry, Graduate School of Science, Osaka University, 1-1 Machikaneyama, Toyonaka, Osaka 560-0043, Japan.
Konoki
Keiichi
K
Matsumori
Nobuaki
N
Oishi
Tohru
T
Murata
Michio
M
D000666:Amphotericin B / Q000737:chemistry*; D002784:Cholesterol / Q000737:chemistry; D004875:Ergosterol / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry*; D008954:Models, Biological; D015394:Molecular Structure; D010713:Phosphatidylcholines / Q000737:chemistry; D013261:Sterols / Q000737:chemistry*; D020349:Surface Plasmon Resonance / Q000379:methods*
D008081:Liposomes; D010713:Phosphatidylcholines; D013261:Sterols; D000666:Amphotericin B; D002784:Cholesterol; C028694:1-palmitoyl-2-oleoylphosphatidylcholine; D004875:Ergosterol