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Query: UNIPROT:P17174 (aspartate aminotransferase)
 14,872 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Three homologous aspartate aminotransferases with virtually identical spatial structures and pairwise amino acid sequence identities of > 40% differ markedly with respect to the yield of renaturation upon dilution from 6 M guanidine hydrochloride (mitochondrial << cytosolic < Escherichia coli). The enzymes also respond differently to molecular chaperones. GroEL/GroES, the Hsp60 homolog of E. coli, increased considerably the yield of renaturation of mitochondrial aspartate aminotransferase and to a lesser extent that of its cytosolic counterpart, but not that of the E. coli enzyme. DnaK/DnaJ/GrpE, the Hsp70 system of E. coli, also increased the yield of renaturation of mitochondrial aspartate aminotransferase. Apparently, specific features in the amino acid sequence or the folding pathway which are independent of the final secondary and tertiary structure determine the interactions of the folding proteins with the chaperone systems.
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PMID:Differential effects of molecular chaperones on refolding of homologous proteins. 854 80

Mitochondrial aspartate aminotransferase is inactivated irreversibly on heating. The inactivated protein aggregates, but aggregation is prevented by the presence of the chaperonin 60 from Escherichia coli (GroEL). The chaperonin increases the rate of thermal inactivation in the temperature range 55-65 degrees C but not at lower temperatures. It has previously been shown [Twomey and Doonan (1997) Biochim. Biophys. Acta 1342, 37-44] that the enzyme switches to a modified, but catalytically active, conformation at approx. 55-60 degrees C and the present results show that this conformation is recognized by and binds to GroEL. The thermally inactivated protein can be released from GroEL in an active form by the addition of chaperonin 10 from E. coli (GroES)/ATP, showing that inactivation is not the result of irreversible chemical changes. These results suggest that the irreversibility of thermal inactivation is due to the formation of an altered conformation with a high kinetic barrier to refolding rather than to any covalent changes. In the absence of chaperonin the unfolded molecules aggregate but this is a consequence, rather than the cause, of irreversible inactivation.
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PMID:Thermal inactivation and chaperonin-mediated renaturation of mitochondrial aspartate aminotransferase. 969 23

At 30 degrees C, the precursor to mitochondrial aspartate aminotransferase (pmAspAT) cannot fold after synthesis in rabbit reticulocyte lysate (RRL), a model for studying intracellular protein folding. However, it folds rapidly once imported into mitochondria. Guanidinium chloride denatured pmAspAT likewise cannot refold at 30 degrees C in a defined in vitro system. However, it refolds rapidly and in good yield in the presence of the intramitochondrial chaperone homologues GroEL and GroES. In this report, we demonstrate that GroEL and GroES can also facilitate the folding of nascent pmAspAT in reticulocyte lysate under conditions where it otherwise would not. When added alone, GroEL arrests the slow folding of nascent pmAspAT and inhibits import into mitochondria. These effects are significantly reversed by adding GroES. These observations suggest that added GroEL participates in an equilibrium with endogenous chaperones in the cytosol which inhibit folding and promote import competence. Native gel electrophoresis suggests that nascent pmAspAT exists in RRL as a heterogeneous population of partially folded species, some of which bind to added GroEL more readily than others. The GroEL-trapped species appear to be among the productive pmAspAT folding intermediates formed in RRL or they at least appear to equilibrate with these intermediates, since they become import competent after GroES-stimulated release from GroEL.
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PMID:The folding of nascent mitochondrial aspartate aminotransferase synthesized in a cell-free extract can be assisted by GroEL and GroES. 1105 Nov 4

The mitochondrial (mAAT) and cytosolic (cAAT) homologous isozymes of aspartate aminotransferase are two relatively large proteins that in their nonnative states interact very differently with GroEL. MgATP alone can increase the rate of GroEL-assisted reactivation of cAAT, yet the presence of GroES is mandatory for mAAT. Addition of an excess of a denatured substrate accelerates reactivation of cAAT in the presence of GroEL, but has no effect on mAAT. These competition studies suggest that the more stringent substrate mAAT forms a thermodynamically stable complex with GroEL, while rebinding affects the slow reactivation kinetics of cAAT with GroEL alone. However, the competitor appears to accelerate the release of cAAT from GroEL, most likely by displacing bound cAAT from the GroEL cavity. Moreover, cAAT, but not mAAT, shows a time-dependent increase in protease resistance while bound to GroEL at low temperature. These results suggest that folding and release of cAAT from GroEL in the absence of cofactors may occur stepwise with certain interactions being broken and reformed until the protein escapes binding. The distinct behavior of these two isozymes most likely results from differences in the structure of the nonnative states that bind to GroEL.
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PMID:Stability and release requirements of the complexes of GroEL with two homologous mammalian aminotransferases. 1123 73

Mitochondrial aspartate aminotransferase (mAspAT) (E.C. 2.6.1.1), an important enzyme in amino acid metabolism, is identical to a fatty acid-binding protein (FABPpm) isolated from plasma membranes of several cell types. Employing a monospecific polyclonal antibody to rat mAspAT, we have used immunogold electron microscopy to study the subcellular distribution of mAspAT in various mammalian tissues. Immunogold labeling of rat tissue sections embedded in LR Gold resin showed strong labeling of mitochondria in all tissues examined (viz. liver, pancreas, pituitary, spleen, heart, kidney, submandibular gland). In addition, strong and specific labeling was also observed at a number of non-mitochondrial sites including various locations in kidney, such as on cell surface in distal tubules and cortical collecting ducts, in condensing vacuoles, along cell boundaries between adjoining cells, and in endothelial cells lining capillaries in the glomerulus. Surface labeling due to mAspAT was also seen in arteriolar endothelial cells and in lymphocytes. These findings support the previous identification of mAspAT as both a mitochondrial enzyme and a plasma membrane protein. It is suggested that in accordance with its established role in other cells and tissues, the surface-located mAspAT in kidney and endothelial cells is involved in the fatty acid transport process. The dual-localization of mAspAT, as well as a large number of other mitochondrial proteins (viz. Hsp60, Hsp10, Cytochrome c, TRAP-1 and P32 (gC1q-R)) in recent studies, within both mitochondria and at various specific extramitochondrial sites raises fundamental questions about the role of mitochondria in cell structure and function, and about the mechanisms that exist in normal cells for protein translocation from mitochondria to other compartments. These results have implications for the role of mitochondria in apoptosis and different diseases.
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PMID:Immunogold localization of mitochondrial aspartate aminotransferase in mitochondria and on the cell surface in normal rat tissues. 1196 39

The interaction of the precursor to mitochondrial aspartate aminotransferase (pmAAT) with GroEL has been studied by electron paramagnetic resonance (EPR) and fluorescence spectroscopy. In the native protein, the spin probe was immobilized when attached to Cys166 at the domain interface, but was fully mobile when introduced at Cys(-19) in the N-terminal presequence peptide. Unfolding of the protein resulted in a highly mobile EPR spectrum for probes introduced at either site. However, the nitroxide group in GroEL-bound pmAAT showed either intermediate or high mobility depending on the spin probe used. Power saturation experiments indicated that the accessibility of the nitroxide side chain to Ni(EDDA) in the GroEL-pmAAT complex was higher than in the native state when in position 166 but lower when at position -19. Similar results were obtained in fluorescence quenching experiments. These data suggest that GroEL binds partly folded states of pmAAT with the presequence peptide probably in direct contact with GroEL. GroES and ATP, but not AMP-PNP or ADP, support refolding of pmAAT. During refolding, the rate of recovery of the native spectroscopic properties of labeled Cys166 is nearly identical to the rate-limiting reactivation step. Thus, correct docking of the large and small domains of pmAAT may be a key structural event in the regain of catalytic activity.
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PMID:Electron paramagnetic resonance and fluorescence studies of the conformation of aspartate aminotransferase bound to GroEL. 1632 39