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Apoptosis is a type of regulated cell death important for normal embryonic development and maintenance of adult tissues by removing excess or dysfunctional cells to ensure proper functioning of organs. The Bcl-2 family of proteins determines whether apoptosis remains suppressed or becomes activated through the balance of interactions among pro-survival and pro-death members. A defining feature of the Bcl-2 family is a BH3 domain that drives interactions between the family members. Isoform 1 of the voltage dependent anion channel (VDAC1) has an important role in metabolism, but was recently found to have high homology with known BH3 domains. This study tested the hypothesis that VDAC1 has a BH3 domain at its N-terminus, making it a novel member of the Bcl-2 family. The ability of the pro-death family member Bax to bind a panel of wild type and mutant VDAC1 peptides was tested to determine if VDAC1 contains a functional BH3 domain, and the residues most important for driving this interaction. The mutant peptides were developed to substitute two or more of the seven hydrophobic residues (designated H0-H5) with those having less hydrophobicity, or replacing a conserved aspartate with arginine. Equilibrium binding of fluorescent-tagged peptides to recombinant Bax was assessed by separating free from bound peptide with spin filters. The two most important residues that promoted VDAC1 interaction with Bax were the conserved aspartic acid, and a hydrophobic proline at ‘H0’. The lower affinity of Bax for VDAC1 compared to the known Bim BH3 domain is likely due to the non-conserved position of the hydrophobic H2 leucine creating steric interference with the close proximity H1 tyrosine. The binding analysis supports the hypothesis that VDAC1 possesses a BH3 domain, although the sequence of the domain is less well suited to interaction with Bax compared to the Bim BH3 domain.
Pearson, Claire, "Is VDAC1 a Novel BCL2 Family Member that Binds BAX?" (2023). Honors Theses. 3021.
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