Interaction of Bombolitin III with Phospholipid Monolayers and Liposomes and Effect on the Activity of Phospholipase A2

Abstract

This study is focused on the characterization of the interaction of the amphiphilic peptide bombolitin III (from the bumblebee Megabombus pennsylvanicus) with phospholipid monolayers and vesicles. It is shown that due to the amphiphilic character of its alpha-helical conformation this water-soluble peptide is able to interact in an ordered fashion with phospholipid organized structures. Depending on the temperature, the subphase, and the particular phosphatidylcholine used, the mixed peptide-phospholipid monolayers can be homogeneous or display phase separation. This behavior was observed by means of the Langmuir film balance technique, coupled with an epifluorescence microscope. In well-defined conditions it is possible to visualize the formation of phase-separated peptide domains at the air-water interface and to study the effect of their presence on the organization of the lipid. The action of phospholipase A2 at the lipid-peptide interface was also followed by means of fluorescence microscopy: some evidence that the enzyme preferentially hydrolyzes the phospholipid that is in contact with the peptide is presented. Furthermore, the presence of bombolitin III in L-alpha-DLPC monolayers causes an increase in the initial speed of degradation with phospholipase A2. These results are in agreement with previous findings that show that the bombolitins are activators in vitro of phospholipase A2. Experiments were also performed with peptide fragments corresponding to the alpha-helical sequences of the protein uteroglobin: despite some amphiphilic character, these peptides do not interact strongly with phospholipid monolayers. Only one of these peptides (corresponding to the helix 4-14 in uteroglobin) is adsorbed in the monolayer in a similar fashion to bombolitin III but does not cause an increase in the activity of phospholipase A2.

Critical pressures in multicomponent lipid monolayers

“…Enzyme activities that are affected by the phase boundaries of lipid domains may provide a means whereby lipid composition itself is regulated through enzyme activity. Although this possibility has not been proposed previously to the best of our knowledge, pub lished experimental work [21][22][23][24][25][26] on the action of en zymes on lipid membranes is not inconsistent with this hypothesis.…”

Interactions of amphotericin B derivative of low toxicity with biological membrane components—the Langmuir monolayer approach

“…The use of the Langmuir method allows for a precise and continuous control of such parameters as molecular packing, physical state, lateral pressure, composition and quality of the surface. In particular, mixed monolayers composed of constituents of biological membranes, such as phospholipids, sterols and glycolipids, provide a highly informative approach for studying intermolecular interactions between membrane components, usually phospholipids, and biomolecules, for example, various drugs [2][3][4][5], hormones [6,7], enzymes [8][9][10][11], proteins [12][13][14][15][16][17][18][19][20][21], etc. Such investigations may be of much help in elucidating a mode of action of many physiological active compounds [22][23][24][25][26][27][28][29].…”

Conformational properties of the amphipathic lytic polypeptide bombolitin II. A circular dichroism, NMR and computer simulation study

“…Because of the high affinity of all bombolitins toward micelles, monolayers and liposomes, it seems resonable to suppose that these peptides influence the activity of PLA, by altering the physical state of the aggregated substrate. This hypothesis was further substantiated by our recent investigation on the interaction of bombolition I11 with phospholipid monolayers and liposomes 30). By the Langmuir balance technique, coupled with epifluorescence microscopy, it was shown that the peptide forms phase-separated peptide domains at the air-water interface, with consequent alteration of the organization of the lipid monolayer.…”