Comparison of the membrane interaction mechanism of two antimicrobial RNases: RNase 3/ECP and RNase 7.
Eosinophil cationic protein (ECP/RNase 3) and the skin derived ribonuclease 7 (RNase 7) are members of the RNase A superfamily. RNase 3 is mainly expressed in eosinophils whereas RNase 7 is primarily secreted by keratinocytes. Both proteins present a broad-spectrum antimicrobial activity and their bactericidal mechanism is dependent on their membrane destabilizing capacities. Using phospholipid vesicles as membrane models, we have characterized the protein membrane association process. Confocal microscopy experiments using giant unilamellar vesicles illustrate the morphological changes of the liposome population. By labelling both lipid bilayers and proteins we have monitored the kinetic of the process. The differential protein ability to release the liposome aqueous content was evaluated together with the micellation and aggregation processes. A distinct morphology of the protein/lipid aggregates was visualized by transmission electron microscopy and the proteins overall secondary structure in a lipid microenvironment was assessed by FTIR. Interestingly, for both RNases the membrane interaction events take place in a different behaviour and timing: RNase 3 triggers first the vesicle aggregation, while RNase 7 induces leakage well before the aggregation step. Their distinct mechanism of action at the membrane level may reflect different in vivo antipathogen functions.
Departament de Bioquímica i Biologia Molecular, Facultat de Biociències, Universitat Autònoma de Barcelona, 08193 Cerdanyola del Vallès, Spain.
Marc
M
Torrent
Daniel
D
Sánchez
Víctor
V
Buzón
M Victòria
MV
Nogués
Josep
J
Cladera
Ester
E
Boix
D000595:Amino Acid Sequence; D055592:Biophysical Phenomena; D047150:Eosinophil Cationic Protein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008081:Liposomes; D018613:Microscopy, Confocal; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012260:Ribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D017550:Spectroscopy, Fourier Transform Infrared
D008081:Liposomes; D046912:Multiprotein Complexes; D011994:Recombinant Proteins; D012260:Ribonucleases; D047150:Eosinophil Cationic Protein; C470000:Ribonuclease 7