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Jason W O'Neill

Publications and source records attributed to Jason W O'Neill.

6 recordsLinked to original sources
PubMed2005

BCL-XL dimerization by three-dimensional domain swapping.

Dimeric interactions among anti- and pro-apoptotic members of the BCL-2 protein family are dynamically regulated and intimately involved in survival and death functions. We report the structure of a BCL-X(L) homodimers a 3D-domain swapped dimer (3DDS). The X-ray crystal structure demonstrates the mutual exchange of carboxy-terminal regions including BH2 (Bcl-2 homology 2) between monomer subunits, with the hinge region occurring at the hairpin turn between the fifth and sixth alpha helices. Both BH3 peptide-binding hydrophobic grooves are unoccupied in the 3DDS dimer and available for BH3 peptide binding, as confirmed by sedimentation velocity analysis. BCL-X(L) 3DDS dimers have increased pore-forming activity compared to monomers, suggesting that 3DDS dimers may act as intermediates in membrane pore formation. Chemical crosslinking studies of Cys-substituted BCL-X(L) proteins demonstrate that 3DDS dimers form in synthetic lipid vesicles.

Cross-Linking Reagents↗
PubMed2004

Mcl-1 is required for Akata6 B-lymphoma cell survival and is converted to a cell death molecule by efficient caspase-mediated cleavage.

Enforced expression of the antiapoptotic Bcl-2 family protein Mcl-1 promotes lymphomagenesis in the mouse; however, the functional role of Mcl-1 in human B-cell lymphoma remains unclear. We demonstrate that Mcl-1 is widely expressed in malignant B-cells, and high-level expression of Mcl-1 is required for B-lymphoma cell survival, since transfection of Mcl-1-specific antisense oligodeoxynucleotides was sufficient to promote apoptosis in Akata6 lymphoma cells. Mcl-1 was efficiently cleaved by caspases at evolutionarily conserved aspartic acid residues in vitro, and during cisplatin-induced apoptosis in B-lymphoma cell lines and spontaneous apoptosis of primary malignant B-cells. Overexpression of the Mcl-1 cleavage product that accumulated during apoptosis was sufficient to kill cells. Therefore, Mcl-1 is an essential survival molecule for B-lymphoma cells and is cleaved by caspases to a death-promoting molecule during apoptosis. In contrast to Mcl-1, Bcl-2 and Bcl-XL were relatively resistant to caspase cleavage in vitro and in intact cells. Interfering with Mcl-1 function appears to be an effective means of inducing apoptosis in Mcl-1-positive B-cell lymphoma, and the unique sensitivity of Mcl-1 to caspase-mediated cleavage suggests an attractive strategy for converting it to a proapoptotic molecule.

Apoptosis↗
PubMed2003

Bcl-XL mutations suppress cellular sensitivity to antimycin A.

Cells expressing high levels of the BCL-X(L) anti-apoptotic protein are preferentially killed by the mitochondrial inhibitor antimycin A (AA). Computational modeling predicts a binding site for AA in the extended hydrophobic groove on BCL-X(L), previously identified as an interface for dimerization to BAX and related proapoptotic proteins. Here, we identify BCL-X(L) hydrophobic groove mutants with normal cellular anti-apoptotic function but suppressed sensitivity to AA. The LD(50) of AA for cells expressing BCL-X(L) mutants directly correlates with the measured in vitro dissociation constants for AA binding. These results indicate that BCL-X(L) is a principal target mediating AA cytotoxicity.

Amino Acid Sequence↗
PubMed2003

Bcl-2-related proteins as drug targets.

The Bcl-2 family of proteins provide the most unambiguous link between mitochondrial functions and apoptosis, as their only (or principal) functions appear to be as regulators of this cell death pathway. Rational drug design to manipulate the functions of these proteins has been hampered by the lack of a clear understanding of a biochemical or molecular function, with disruption of intra-family protein-protein interactions as the only known, but daunting, objective. There has been substantial progress in this task using molecular modeling and drug leads. The prospects are also good for development of chemical tools for functional analysis of the Bcl-2 proteins.

Animals↗
PubMed2002

Accurate computer-based design of a new backbone conformation in the second turn of protein L.

The rational design of loops and turns is a key step towards creating proteins with new functions. We used a computational design procedure to create new backbone conformations in the second turn of protein L. The Protein Data Bank was searched for alternative turn conformations, and sequences optimal for these turns in the context of protein L were identified using a Monte Carlo search procedure and an energy function that favors close packing. Two variants containing 12 and 14 mutations were found to be as stable as wild-type protein L. The crystal structure of one of the variants has been solved at a resolution of 1.9 A, and the backbone conformation in the second turn is remarkably close to that of the in silico model (1.1 A RMSD) while it differs significantly from that of wild-type protein L (the turn residues are displaced by an average of 7.2 A). The folding rates of the redesigned proteins are greater than that of the wild-type protein and in contrast to wild-type protein L the second beta-turn appears to be formed at the rate limiting step in folding.

Amino Acid Sequence↗