Measuring pore formation by Bcl-2 family proteins.
Two methods for assaying Bcl-2 protein family-induced solute efflux from liposomes have been outlined. They utilize either ion-selective electrodes to follow ion efflux or fluorescence to monitor changes in fluorescence of the liposome-encapsulated dye SPQ or carboxyfluorescein. Both methods provide a simple means of determining protein activity. These methods do not have the capability to detect either single-channel conductivity or ion selectivity, but they indicate whether the bulk of the protein population is inducing solute efflux. Although in in vivo significance of Bcl-2 protein family pore formation remains to be determined, in vitro measurements of channel activity should provide a means to determine whether a given protein preparation has activity and whether mutations have an adverse effect on channel formation.
Schendel
S L
SL
Burnham Institute, La Jolla, California 92037, USA.
Reed
J C
JC
D000818:Animals; D017209:Apoptosis; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods; D005452:Fluoresceins; D005456:Fluorescent Dyes; D008081:Liposomes; D011487:Protein Conformation; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry* / Q000378:metabolism*; D011806:Quinolinium Compounds; D013050:Spectrometry, Fluorescence / Q000379:methods; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein
D005452:Fluoresceins; D005456:Fluorescent Dyes; D008081:Liposomes; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011806:Quinolinium Compounds; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; C024098:6-carboxyfluorescein; C052335:6-methoxy-N-(3-sulfopropyl)quinolinium