Monitoring the looping up of acyl chain labeled NBD lipids in membranes as a function of membrane phase state

Abstract

Lipids that are labeled with the NBD (7-nitrobenz-2-oxa-1,3-diazol-4-yl) group are widely used as fluorescent analogues of native lipids in biological and model membranes to monitor a variety of processes. The NBD group of acyl chain labeled NBD lipids is known to loop up to the membrane interface in fluid phase membranes. However, the organization of these lipids in gel phase membranes is not resolved. In this paper, we monitored the influence of the membrane phase state on the looping up behavior of acyl chain labeled NBD lipids utilizing red edge excitation shift (REES) and other sensitive fluorescence approaches. Interestingly, our REES results indicate that NBD group of lipids, which are labeled at the fatty acyl region, resides in the more hydrophobic region in gel phase membranes, and complete looping of the NBD group occurs only in the fluid phase. This is supported by other fluorescence parameters such as polarization and lifetime. Taken together, our results demonstrate that membrane packing, which depends on temperature and the phase state of the membrane, significantly affects the localization of acyl chain labeled NBD lipids. In view of the wide ranging use of NBD-labeled lipids in cell and membrane biology, these results could have potentially important implications in future studies involving these lipids as tracers.

Detection of Lipid Domains in Model and Cell Membranes by Fluorescence Lifetime Imaging Microscopy of Fluorescent Lipid Analogues

“…In contrast, it was shifted to ϳ12 ns in lo domains ( 2(lo) ) reflecting the higher degree of order in those domains. In line with previous results (40,42) we surmise that the relocation dynamics of the NBD moiety is sensitive to lipid packing and that the increase in the degree of order when going from a ld to a lo phase causes the large shift of 2 toward longer lifetimes. We detected only minor amounts if at all of 2(lo) in the ld domain and no contributions of 2(ld) in the lo domain.…”

Miltefosine and BODIPY-labeled alkylphosphocholine with leishmanicidal activity: Aggregation properties and interaction with model membranes

“…The NBD group of sn-2 acyl chain NBD-labeled phosphatidylcholine (NBD-PC) may loop to a location near the lipid/water interface of phospholipid bilayers and in C6-NBD-PC it does not penetrate the bilayer beyond the C4-C5 atoms of sn-2 chains of the host lipid (45). The probes are able to monitor structural changes in lipid bilayers like those occurring in thermal phase transition of lipid vesicles (46,47).…”

“…In vesicles of DPPC in the fluid phase, the NBD group in the probe C6-NBD-PC is located in the region of the glycerol backbone and it loops to a transverse location closer to the interface (53). Below the phase transition T m , the NBD group in NBD-PC may localize in deeper regions of the bilayer, with a distribution along the membrane axis (47).…”

Anisotropia de fluorescência: aplicações em membranas modelo.

“…Ainda não está bem elucidado o comportamento da molécula ligada nas diferentes posições dos fosfolipídios inseridos nas membranas lipídicas [49], principalmente na fase gel, porém entende-se que o grupo NBD tem preferência pela região da interface da membrana, tanto na situação em que se encontra ligado aos carbonos de uma das cadeias de ácido graxo da região hidrofóbica, como o C6-NBD-PC e o C12-NBD-PC, como ligado à cabeça polar do fosfolipídio, como o NBD-PE [10]. Abaixo estão representadas diferentes localizações do NBD nas moléculas fosfolipídicas.…”

“…Este deslocamento espectral indica aumento na polaridade onde o DPH está localizado [28]. A mudança na polaridade acontece devido à penetração de moléculas de água da suspensão na vesícula fosfolipídica, que se torna possível devido à desordem dos fosfolipídios promovida por maior agitação térmica na fase fluida [49]. …”