title: Identification of Hsc70 binding sites in mitochondrial aspartate aminotransferase.
abstract: Hsc70 binds acid-unfolded mitochondrial aspartate aminotransferase (mAAT), forming either soluble or insoluble complexes depending on the relative concentrations of the proteins. Using partial proteolysis of Hsc70-mAAT complexes in combination with MALDI-TOF mass spectrometry, we have identified several potential Hsc70-binding regions in the mAAT polypeptide. Only one mAAT peptide was found bound to Hsc70 in the insoluble complexes while nine peptides arising from eight sequence regions of mAAT were found associated with Hsc70 in the soluble complexes. Most of these binding sites map to secondary structure elements, particularly alpha-helix, that are partly exposed on the surface of the folded structure. These results suggest that these peptide regions must not only be exposed but still in a flexible extended conformation in the mAAT folding intermediates recognized by Hsc70. Thus, for mAAT the discrimination between native and non-native structures by Hsc70 may rely more on the level of structure of the binding sites than on their degree of exposure to the solvent in the native structure.
authors.affiliation: Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri-Kansas City, 64110-2499, USA.
authors.forename: Antonio
authors.initials: A
authors.lastname: Artigues
authors.forename: Ana
authors.initials: A
authors.lastname: Iriarte
authors.forename: Marino
authors.initials: M
authors.lastname: Martinez-Carrion
mesh_terms: D000818:Animals; D024442:Aspartate Aminotransferase, Mitochondrial / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D050883:HSC70 Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017510:Protein Folding; D017433:Protein Structure, Secondary; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D050883:HSC70 Heat-Shock Proteins; C494372:Hspa8 protein, mouse; D046912:Multiprotein Complexes; D010455:Peptides; D024442:Aspartate Aminotransferase, Mitochondrial