Reconstituted proteolipid vesicles prepared from Mycoplasma fermentans membranes are able to bind and fuse with Molt-3 cells.
We describe and characterize reconstituted proteolipid vesicles (rPLV) prepared from solubilized Mycoplasma fermentans membranes and studied their binding to and fusion with host Molt-3 cells. The rPLV were prepared following membrane solubilization by Triton X-100 and detergent removal by SM-2 resin beads. The vesicles thus obtained had a rather uniform diameter of about 1 microm and were sealed as monitored by measuring in an assay that measures the quenching by sodium dithionite of a hydrophobic fluorescent probe incorporated into the rPLV membrane. The rPLV adhered to Molt-3 cells and, based on measurements of lipid mixing, fused with the host cells at a similar rate and to about the same extent as intact M. fermentans. Preliminary experiments showed that a chimeric protein, GnRH-PE66, could be encapsulated within these rPLV, opening the way to develop a system for the transfer of high-molecular weight soluble molecules, encapsulated in the rPLV, to target eukaryotic cells.
Rechnitzer
Hagai
H
Department of Membrane and Ultrastructure Research, The Hebrew University--Hadassah Medical School, P.O. Box 12272, Jerusalem, 91120, Israel.
Rottem
Shlomo
S
D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry*; D005098:Exotoxins / Q000032:analysis; D005456:Fluorescent Dyes; D007987:Gonadotropin-Releasing Hormone / Q000032:analysis; D006801:Humans; D008081:Liposomes / Q000737:chemistry*; D008561:Membrane Fusion; D016858:Mycoplasma fermentans / Q000648:ultrastructure*; D011510:Proteolipids / Q000737:chemistry*; D011549:Pseudomonas / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000032:analysis
D005098:Exotoxins; D005456:Fluorescent Dyes; D008081:Liposomes; D011510:Proteolipids; D011993:Recombinant Fusion Proteins; D007987:Gonadotropin-Releasing Hormone