title: A membrane-targeted BID BCL-2 homology 3 peptide is sufficient for high potency activation of BAX in vitro.
abstract: The multidomain pro-apoptotic proteins BAX and BAK constitute an essential gateway to mitochondrial dysfunction and programmed cell death. Among the "BCL-2 homology (BH) 3-only" members of pro-apoptotic proteins, truncated BID (tBID) has been implicated in direct BAX activation, although an explicit molecular mechanism remains elusive. We find that BID BH3 peptide alone at submicromolar concentrations cannot activate BAX or complement BID BH3 mutant-tBID in mitochondrial and liposomal release assays. Because tBID contains structurally defined membrane association domains, we investigated whether membrane targeting of BID BH3 peptide would be sufficient to restore its pro-apoptotic activity. We developed a Ni(2+)-nitrilotriacetic acid liposomal assay system that efficiently conjugates histidine-tagged peptides to a simulated outer mitochondrial membrane surface. Strikingly, nanomolar concentrations of a synthetic BID BH3 peptide that is chemically tethered to the liposomal membrane activated BAX almost as efficiently as tBID itself. These results highlight the importance of membrane targeting of the BID BH3 domain in tBID-mediated BAX activation and support a model in which tBID engages BAX to trigger its pro-apoptotic activity.
authors.affiliation: Howard Hughes Medical Institute, the Department of Pathology and Medicine, Harvard Medical School, Dana-Farber Cancer Institute, Boston, MA 02115, USA. kyoung_joon_oh@dfci.harvard.edu
authors.forename: Kyoung Joon
authors.initials: KJ
authors.lastname: Oh
authors.forename: Scott
authors.initials: S
authors.lastname: Barbuto
authors.forename: Kenneth
authors.initials: K
authors.lastname: Pitter
authors.forename: Joel
authors.initials: J
authors.lastname: Morash
authors.forename: Loren D
authors.initials: LD
authors.lastname: Walensky
authors.forename: Stanley J
authors.initials: SJ
authors.lastname: Korsmeyer
mesh_terms: D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein / Q000737:chemistry / Q000502:physiology*; D006639:Histidine / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D008954:Models, Biological; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry; D051028:bcl-2-Associated X Protein / Q000502:physiology*; D051020:bcl-X Protein / Q000378:metabolism
chemical_list: D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; D008055:Lipids; D008081:Liposomes; D010455:Peptides; D019253:Proto-Oncogene Proteins c-bcl-2; D011994:Recombinant Proteins; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D006639:Histidine