Oligomeric Aip2p/Dld2p modifies the protein conformation of both properly folded and misfolded substrates in vitro

Oligomeric actin-interacting protein 2 (Aip2p) [Nat. Struct. Biol. 2 (1995) 28]/D-lactate dehydrogenase protein 2 (Dld2p) [Yeast 15 (1999) 1377, Biochem. Biophys. Res. Commun. 295 (2002) 910] exhibits the unique grapple-like structure with an ATP-dependent opening [Biochem. Biophys. Res. Commun. 320 (2004) 1271], which is required for the F-actin conformation modifying activity in vitro and in vivo [Biochem. Biophys. Res. Commun. 319 (2004) 78]. To further investigate the molecular nature of oligomeric Aip2p/Dld2p, the substrate specificity of its binding and protein conformation modifying activity was examined. In the presence of 1mM ATP or AMP-PNP, oligomeric Aip2p/Dld2p bound to all substrates so far examined, and modified the conformation of actin, DNase I, the mature form of invertase, prepro-alpha-factor, pro-alpha-factor, and mitochondrial superoxide dismutase, as determined by the trypsin susceptibility assay. Of note, the activity could modify even the conformation of pathogenic highly aggregated polypeptides, such as recombinant prion protein in beta-sheet form, alpha-synuclein, and amyloid beta (1-42) in the presence of ATP. The in vivo protein conformation modifying activity, however, depends on the growth stage; the most significant substrate modification activity was observed in yeast cells at the log phase, suggesting the presence of a cofactor/s in yeast cells, where F-actin is supposed to be a major target in vivo. These data further support our previous notion that the oligomeric Aip2p/Dld2p may belong to an unusual class of molecular chaperones [Biochem. Biophys. Res. Commun. 320 (2004) 1271], which can target both properly folded and misfolded proteins in an ATP-dependent manner in vitro.