title: Membrane fusion mediated by phospholipase C under endosomal pH conditions.
abstract: Phospholipase C (PLC) is considered to be one of key enzymes for the design of drug delivery system using the endocytosis route, because PLC can catalyze the membrane fusion between cell membranes and phospholipid vehicles (liposomes). Membrane fusion by PLC was then studied under various pHs to model the endosomal environment. The used liposomes were composed of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), and cholesterol (Ch). The membrane fusion was dominated by the enzymatic reaction at pH 6-7.5. In contrast, the membrane perturbation effect due to the conformational change of PLC could induce the membrane fusion at around pH 4. The maximal value of membrane fusion was observed at around pH 5 for three liposomes in the order of DOPC<DOPC/DPPC (1:1)<DOPC/DPPC/Ch (1:1:1). From the experiments with the hydrophobic fluorescence probe and the circular dichroic method, it was revealed that PLC took a molten-globule state, with a large fluctuation and an enzymatic activity, at around pH 4-5. The DAG-rich domain enzymatically produced by PLC played role for the field for the membrane perturbation enough to lead to the membrane fusion. The present finding would be helpful to understand the behavior of membrane fusion under the late endosomal pH condition in cell system.
authors.affiliation: Department of Chemical Science and Engineering, Graduate School of Engineering Science, Osaka University, 1-3 Machikaneyama-cho, Toyonaka, Osaka 560-8531, Japan.
authors.forename: Toshinori
authors.initials: T
authors.lastname: Shimanouchi
authors.forename: Hidenori
authors.initials: H
authors.lastname: Kawasaki
authors.forename: Makoto
authors.initials: M
authors.lastname: Fuse
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Umakoshi
authors.forename: Ryoichi
authors.initials: R
authors.lastname: Kuboi
mesh_terms: D015060:1,2-Dipalmitoylphosphatidylcholine / Q000737:chemistry; D001409:Bacillus cereus / Q000201:enzymology*; D002784:Cholesterol / Q000737:chemistry; D002942:Circular Dichroism; D011992:Endosomes / Q000378:metabolism*; D005452:Fluoresceins / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D008081:Liposomes / Q000737:chemistry; D008561:Membrane Fusion; D010713:Phosphatidylcholines / Q000737:chemistry; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence; D010738:Type C Phospholipases / Q000737:chemistry / Q000378:metabolism*
chemical_list: D005452:Fluoresceins; D008024:Ligands; D008081:Liposomes; D010713:Phosphatidylcholines; D015060:1,2-Dipalmitoylphosphatidylcholine; D002784:Cholesterol; D010738:Type C Phospholipases; C017251:1,2-oleoylphosphatidylcholine; C007740:fluorexon