title: Alpha-helical destabilization of the Bcl-2-BH4-domain peptide abolishes its ability to inhibit the IP3 receptor.
abstract: The anti-apoptotic Bcl-2 protein is the founding member and namesake of the Bcl-2-protein family. It has recently been demonstrated that Bcl-2, apart from its anti-apoptotic role at mitochondrial membranes, can also directly interact with the inositol 1,4,5-trisphosphate receptor (IP3R), the primary Ca(2+)-release channel in the endoplasmic reticulum (ER). Bcl-2 can thereby reduce pro-apoptotic IP3R-mediated Ca(2+) release from the ER. Moreover, the Bcl-2 homology domain 4 (Bcl-2-BH4) has been identified as essential and sufficient for this IP3R-mediated anti-apoptotic activity. In the present study, we investigated whether the reported inhibitory effect of a Bcl-2-BH4 peptide on the IP 3R1 was related to the distinctive α-helical conformation of the BH4 domain peptide. We therefore designed a peptide with two glycine "hinges" replacing residues I14 and V15, of the wild-type Bcl-2-BH4 domain (Bcl-2-BH4-IV/GG). By comparing the structural and functional properties of the Bcl-2-BH4-IV/GG peptide with its native counterpart, we found that the variant contained reduced α-helicity, neither bound nor inhibited the IP 3R1 channel, and in turn lost its anti-apoptotic effect. Similar results were obtained with other substitutions in Bcl-2-BH4 that destabilized the α-helix with concomitant loss of IP3R inhibition. These results provide new insights for the further development of Bcl-2-BH4-derived peptides as specific inhibitors of the IP3R with significant pharmacological implications.
authors.affiliation: Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine, Leuven, Belgium.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Monaco
authors.forename: Elke
authors.initials: E
authors.lastname: Decrock
authors.forename: Koen
authors.initials: K
authors.lastname: Nuyts
authors.forename: Larry E
authors.initials: LE
authors.lastname: Wagner
authors.forename: Tomas
authors.initials: T
authors.lastname: Luyten
authors.forename: Sergei V
authors.initials: SV
authors.lastname: Strelkov
authors.forename: Ludwig
authors.initials: L
authors.lastname: Missiaen
authors.forename: Wim M
authors.initials: WM
authors.lastname: De Borggraeve
authors.forename: Luc
authors.initials: L
authors.lastname: Leybaert
authors.forename: David I
authors.initials: DI
authors.lastname: Yule
authors.forename: Humbert
authors.initials: H
authors.lastname: De Smedt
authors.forename: Jan B
authors.initials: JB
authors.lastname: Parys
authors.forename: Geert
authors.initials: G
authors.lastname: Bultynck
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D045744:Cell Line, Tumor; D057927:Hydrophobic and Hydrophilic Interactions; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000037:antagonists & inhibitors*; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D010539:Permeability / Q000187:drug effects; D055550:Protein Stability; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry*
chemical_list: D053496:Inositol 1,4,5-Trisphosphate Receptors; D010446:Peptide Fragments; D019253:Proto-Oncogene Proteins c-bcl-2; D002118:Calcium