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UNIVERSITY OF BUCHAREST
FACULTY OF PHYSICS Guest
2024-11-23 17:53 |
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Conference: Bucharest University Faculty of Physics 2009 Meeting
Section: Biophysics; Medical Physics
Title:
Fluorescence techniques used to prove AMP’s (antimicrobial peptides) insertion into lipid vesicles
Authors:
Mihaela Bacalum, Mihai Radu
Affiliation:
“Horia Hulubei” National Institute for Physics and Nuclear Engineering, Department of Health and Environmental Physics, Atomistilor 407, Magurele, 077125, Romania
E-mail
bmihaela@nipne.ro
Keywords:
Abstract:
Antimicrobial peptides (AMPs) are small peptides produced by different living organisms. Due to their antimicrobial activity they are used alone or with antibiotics against bacteria, viruses and fungi. Due to their specific structure, the AMP’s are attaching very easily to lipid membranes by inserting into them and forming transmembrane pores. For a better understanding of AMP’s action mechanisms we performed experiments on model membranes (e.g., liposomes) using fluorescence techniques. AMP’s used in our experiment were: Gramicidin A, Gramicidin C, Melittin and Indolicidin. Liposomes were prepared from DPhPC and from a mixture of DPhPC and a fluorescently labeled lipid βDPH-HPC (in different concentration.In both cases the final concentration of lipids was 78 μM. AMP’s insertion into the lipid bilayer was first proved by the shift in the tryptophan fluorescence maximum, from ~ 350 nm (value characteristic to tryptophan in a hydrophilic environment – phosphate buffer saline) to ~ 335 nm (peptide found in a more hydrophobic environment – inserted in the lipid bilayer). Fluorescence resonance energy transfer (FRET)was also used to prove the AMPs insertion. In this case, tryptophan residues were the donor, and βDPH-HPC (a fluorescent lipid) was the acceptor. After processing the data, we observed a decrease in tryptophan fluorescence and an increase in βDPH-HPC fluorescence, when the concentration of βDPH-HPC was increasing. Fluorescence anisotropy measurements were then used to observe any modifications in lipid mobility when these peptides are inserting into lipid membranes.
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