STRUCTURAL FEATURES OF THE PRECURSOR TO MITOCHONDRIAL ASPARTATE-AMINOTRANSFERASE RESPONSIBLE FOR BINDING TO HSP70

The precursor (pmAspAT) and mature (mAspAT) forms of mitochondrial asp artate aminotransferase interact with hsp70 very early during translat ion when synthesized in either rabbit reticulocyte lysate or wheat ger m extract (Lain, B., Iriarte, A., and Martinez-Carrion, (1994) J, Biol , Chem, 269, 15588-15596). The nature of the structural elements respo nsible for recognition and binding of this protein to hsp70 has been s tudied by examining the folding and potential association with the cha perone of several engineered forms of this enzyme, Whereas pmAspAT and mAspAT bind hsp70 very early during translation, the cytosolic form o f this enzyme (cAspAT) does not interact with hsp70. A fusion protein consisting of the mitochondrial presequence peptide attached to the am ino terminus of cAspAT associates with hsp70 only after the protein ha s acquired its native-like conformation, apparently through binding to the presequence exposed on the surface of the folded protein. Deletio n of the amino-terminal segment of mAspAT or its replacement with the corresponding domain from the cytosolic isozyme eliminates the cotrans lational binding of hsp70 to the mitochondrial protein, We conclude th at both the presequence and NH2-terminal region of pmAspAT represent r ecognition signals for binding of hsp79 to the newly synthesized mitoc hondrial precursor. Results from competition studies with synthetic pe ptides support this conclusion. The ability of hsp70 to discriminate b etween these two highly homologous proteins probably involves the reco gnition of specific sequence elements in the NH2-terminal portion of t he mitochondrial protein and may relate to their separate localization in the cell. A slower folding rate and higher affinity for cytosolic chaperones may represent evolutionary adaptations of translocated mito chondrial proteins to ensure their efficient importation into the orga nelle.