Apoptosis is initiated when Bcl-2 and its prosurvival relatives are engaged by proapoptotic BH3-only proteins via interaction of its BH3 domain with a groove on the Bcl-2-like proteins. These interactions have been considered promiscuous, but our analysis of the affinity of eight BH3 peptides for five Bcl-2-like proteins has revealed that the interactions vary over 10,000-fold in affinity, and accordingly, only certain protein pairs associate inside cells. Bim and Puma potently engaged all the prosurvival proteins comparably. Bad, however, bound tightly to Bcl-2, Bcl-xL, and Bcl-w but only weakly to A1 and not to Mcl-1. Strikingly, Noxa bound only Mcl-1 and A1. In accord with their complementary binding, Bad and Noxa cooperated to induce potent killing. The results suggest that apoptosis relies on selective interactions between particular subsets of these proteins and that it should be feasible to discover BH3-mimetic drugs that inactivate specific prosurvival targets.

Apoptosis is held in check by prosurvival proteins of the Bcl-2 family. The distantly related BH3-only proteins bind to and antagonize them, thereby promoting apoptosis. Whereas binding of the BH3-only protein Noxa to prosurvival Mcl-1 induces Mcl-1 degradation by the proteasome, binding of another BH3-only ligand, Bim, elevates Mcl-1 protein levels. We compared the threedimensional structures of the complexes formed between BH3 peptides of both Bim and Noxa, and we show that a discrete C-terminal sequence of the Noxa BH3 is necessary to instigate Mcl-1 degradation.apoptosis ͉ Bim ͉ Noxa ͉ crystallography T he mammalian Bcl-2-related antiapoptotic proteins (Bcl-2, Bcl-x L , Bcl-w, Mcl-1, and A1) are critical for maintaining cell survival during development or in response to various stress stimuli (1). They share up to four Bcl-2 homology domains, BH1 through BH4, and contain a putative membrane anchoring sequence at their C termini. Structural studies on proteins lacking only this C-terminal segment reveal that the Bcl-2 family fold is that of an all-helical protein in which the BH1, BH2, and BH3 domains are spatially clustered around a depression on the protein surface (2-5). In response to death signals, such as cytotoxic agents or radiation, a related protein family (the BH3-only proteins) antagonizes the function of the antiapoptotic proteins. The BH3 domains of these proapoptotic molecules form an amphipathic ␣-helical fold when bound to a groove lined by the BH1, BH2, and BH3 domains of antiapoptotic proteins such as Bcl-x L (6-8), a step thought to be important for apoptosis induction.Mcl-1 (myeloid cell factor 1) (9) has features distinguishing it from the other prosurvival proteins. It has a central and nonredundant role in the maintenance of progenitor and stem cells (10-12). The levels of Mcl-1 are highly regulated. In some cell types, signals for differentiation trigger its up-regulation (13), whereas basal levels are controlled, at least in part, by the ubiquitin-proteasome machinery. The HECT domain-containing E3 ligase Mule controls basal Mcl-1 protein abundance and induces its degradation in response to DNA-damaging agents such as cisplatin (14). Mule harbors a BH3 domain that allows it to bind Mcl-1. Noxa is a BH3-only protein that can bind and trigger proteasome-mediated Mcl-1 degradation (15). Whether Mule and Noxa contribute to the proteasomal degradation of Mcl-1 in response to UV irradiation (16) or viral infection is unclear (17). Furthermore, the structural basis for Mcl-1 degradation induced by Noxa is unknown.Recently we have shown that the five mammalian antiapoptotic molecules cluster into two classes; one (containing Bcl-2, Bcl-x L , and Bcl-w) is neutralized by the BH3-only protein Bad, and the other (containing Mcl-1 and A1) is neutralized by the BH3-only protein Noxa (18). Inactivation of both subsets of prosurvival proteins appears necessary for cell death to proceed. Interestingly, the recently described Bcl-2 antagonist ABT-737 (19) is a Bad-like BH3 mimetic and does not bind M...

A major source of free radical production in the brain derives from copper. To prevent metal-mediated oxidative stress, cells have evolved complex metal transport systems. The Alzheimer's disease amyloid precursor protein (APP) is a major regulator of neuronal copper homeostasis. APP knockout mice have elevated copper levels in the cerebral cortex, whereas APP-overexpressing transgenic mice have reduced brain copper levels. Importantly, copper binding to APP can greatly reduce amyloid ␤ production in vitro. To understand this interaction at the molecular level we solved the structure of the APP copper binding domain (CuBD) and found that it contains a novel copper binding site that favors Cu(I) coordination. The surface location of this site, structural homology of CuBD to copper chaperones, and the role of APP in neuronal copper homeostasis are consistent with the CuBD acting as a neuronal metallotransporter.

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