title: Nuclear magnetic resonance study of protein-protein interactions involving apoptosis regulator Diva (Boo) and the BH3 domain of proapoptotic Bcl-2 members.
abstract: According to biochemical assays, the Bcl-2 protein Diva from mouse regulates programmed cell death by heterodimerizing with other members of the family and by interacting with the apoptotic protease-activating factor Apaf-1. In typical Bcl-2 heterodimers, peptide fragments comprising the Bcl-2 homology domain 3 (BH3 domain) of proapoptotic members are capable of forming functional complexes with prosurvival proteins. High-resolution structural studies have revealed that the BH3 peptide forms an α-helix positioned in a canonical hydrophobic cleft of the antiapoptotic protein. Because Diva shows mutations in conserved residues within this area, it has been proposed to have a different interacting surface. However, we showed previously that Diva binds through the canonical groove the BH3 peptide of the human Bcl-2 killing member Harakiri. To further test Diva's binding capabilities, here we show Nuclear Magnetic Resonance (NMR) data, indicating that Diva binds peptides derived from the BH3 domain of several other proapoptotic Bcl-2 proteins, including mouse Harakiri, Bid, Bak and Bmf. We have measured the binding affinities of the heterodimers, which show significant variability. Structural models of the protein-peptide complexes based on NMR chemical shift perturbation data indicate that the binding surface is analogous. These models do not rely on NMR NOE (Nuclear Overhauser Effect) data, and thus our results can only suggest that the complexes share similar intermolecular interactions. However, the observed affinity differences correlate with the α-helical population of the BH3-peptides obtained from circular dichroism experiments, which highlights a role of conformational selection in the binding mechanism. Altogether, our results shed light on important factors governing Diva-BH3 peptide molecular recognition mode.
authors.affiliation: Chemical and Physical Biology Department, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Ramiro de Maeztu, 9, Madrid, 28040, Spain.
authors.forename: Clara M
authors.initials: CM
authors.lastname: Santiveri
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Sborgi
authors.forename: Eva
authors.initials: E
authors.lastname: de Alba
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000502:physiology; D051017:Apoptosis Regulatory Proteins / Q000737:chemistry / Q000378:metabolism; D051023:BH3 Interacting Domain Death Agonist Protein / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs / Q000502:physiology; D060066:Protein Interaction Maps; D017433:Protein Structure, Secondary; D011518:Proto-Oncogene Proteins / Q000737:chemistry; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000737:chemistry* / Q000378:metabolism*; D050998:bcl-2 Homologous Antagonist-Killer Protein / Q000737:chemistry / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; C494772:BAK1 protein, human; C117144:BCL2-like 10 protein; D051023:BH3 Interacting Domain Death Agonist Protein; C494814:BID protein, human; C486655:BMF protein, human; C474007:Bax protein (53-86); C484586:HRK protein, human; D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein