Two forms of the apoptosis-linked protein ALG-2 with different Ca2+ affinities and target recognition

The apoptosis-linked gene ALG-2 encodes a Ca2+-binding protein of the penta EF-hand family, To investigate the Ca2+ binding properties of the recombin ant ALG-2 protein, Re have cloned ALG-2 cDNA from mouse liver mRNA. Sequenc e analysis showed that two types of clones were present. One (named ALG-2,5 ) corresponds to the published ALG-2 sequence (Vito, P., Lacana, E., and D' Adamio, L. (1996) Science 271, 521-525); the second (named ALG-2,1) is 6 nu cleotides shorter, and the corresponding protein lacks the amino acid resid ues Gly(121) and Phe(122), Both transcripts are present in mouse tissues in the same 2:1 molar ratio, The ALG-2,5 and ALG-2,1 recombinant proteins are fully soluble in the metal-free form but can be precipitated from bacteria l lysates by Ca2+. In the presence of TR een the Ca2+ binding profiles disp lay two high affinity sites with [Ca2+](0.5) values of 1.2 and 3.1 mu M for ALG-2,5 and ALG-2,1, respectively, plus one low affinity site. Using the y east two-hybrid system we demonstrate that both proteins have a strong tend ency to form homo- and heterodimers. In contrast to ALG-2,5, the ALG-2,1 is oform does not interact with the target protein AIP-1, earlier described to play a role in apoptosis (Vito, P., Pellegrini, L., Guiet, C., and D'Adami o, L. (1999) J. Biol, Chem. 274, 1533-1540), We propose that the minor sequ ence difference between ALG-2,5 and ALG-2,1 affects the Ca2+ binding proper ties and function of the proteins.