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

The primary structure of tyrosine aminotransferase, as deduced from the nucleotide sequence of complementary DNA, was confirmed by fast atom bombardment mass spectrometry of tryptic peptides derived from the purified protein. Limited digestion of the native enzyme with trypsin released an acetylated, amino-terminal peptide; the new amino terminus in the modified enzyme was Val65. Endogenous proteases generated a chromatographically separable form of tyrosine aminotransferase that began at Lys35. Neither trypsin nor the other proteases altered the catalytic activity of tyrosine aminotransferase. Reduction of the holoenzyme with sodium borohydride yielded a major tryptic peptide containing phosphopyridoxamine bound to lysine 280, which probably functions in transamination. The carboxyl terminus of tyrosine aminotransferase contains features that typify proteins with short half-lives; it includes two negatively charged, hydrophilic segments that are enriched for glutamyl residues and are similar to a PEST region in ornithine decarboxylase (Rogers, S., Wells, R., and Rechsteiner, M. (1986) Science 234, 364-368). Tyrosine aminotransferase belongs to a superfamily of enzymes which includes aspartate aminotransferase and can be aligned so that many invariant, functional residues coincide. Like the isoenzymes of aspartate aminotransferase, tyrosine aminotransferase may contain two domains, with a central, catalytic core, and a small domain made up of both amino- and carboxyl-terminal components. We speculate that the exposed small domain may confer the unusually rapid degradative rate that characterizes this enzyme.
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PMID:The structure of tyrosine aminotransferase. Evidence for domains involved in catalysis and enzyme turnover. 256 40

The effect of various proteases (trypsin, chymotrypsin, subtilisin, protease 401, and thermolysin) on the mitochondrial isoenzyme (m-AST) and cytoplasmic isoenzyme (c-AST) of human and swine aspartate aminotransferase (AST;EC 2.6.1.1) was evaluated. All procedures including the reaction with proteases and the subsequent determination of the AST activity were carried out in an automatic analyzer. The mammalian c-AST was efficiently inactivated by chymotrypsin, subtilisin and protease 401 while m-AST activity decreased very slowly with these proteases. Thermolysin and trypsin showed much less effect on c-AST activity. Especially, chymotrypsin at concentrations of 0.5-1.0 g/L inactivated human c-AST almost completely but showed no detectable inactivating effect on m-AST. Thus chymotrypsin appears to be the most suitable protease for the differential determination of AST isoenzymes in human serum. Further studies on the effects of proteases with AST from other species showed that Escherichia coli AST resembled mammalian m-AST while Pseudomonas AST resembled c-AST.
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PMID:Determination of human aspartate aminotransferase isoenzymes by their differential sensitivity to proteases. 306 46

Genetic engineering is a powerful tool for exploring correlations between structure and function in proteins, but as yet we are unable to use it for effective protein design. One of the most interesting examples, which would seem to be obvious, is reversing the polarity of an ion pair. Changing a positively charged protein group, that provides a strong binding for negative substrates, to a negative group is expected to provide an effective binding site for a positively charged substrate. But several recent experiments on aspartate aminotransferase, trypsin and aspartate transcarbamoylase (Schachman, H. K. personal communication) have indicated that polarity reversal is not so successful. Here we argue that the same factors that make the enzyme an effective system for the (-+) pair will make it a much less effective system for the (+-) pair. We also point out that the unusually low effective dielectric constant (epsilon approximately equal to 13) for the (-+) interaction is due to its microenvironment and this will destabilize a (+-) arrangement having an entirely different dielectric constant (epsilon approximately equal to 80). The calculations presented here evaluate the energetics of ion pairs in protein active sites on a semiquantitative level. This is particularly important when dealing with strong, functionally important interactions that are difficult to evaluate with macroscopic models.
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PMID:Why ion pair reversal by protein engineering is unlikely to succeed. 316 61

Treatment of mitochondrial aspartate aminotransferase from rat liver with trypsin leads to specific cleavage of the bonds between residues 26 and 27, and residues 31 and 32. The proteolysed enzyme has only a small residual catalytic activity, but retains a conformation similar to that of the native form as judged by accessibility and reactivity of cysteine residues. Proteolysis abolishes the ability of the enzyme either to bind to mitochondria or to be imported into the organelles. This suggests that the N-terminal segment of the native enzyme is essential for both of these functions, at least in the model system used to study the import process.
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PMID:Removal of an N-terminal peptide from mitochondrial aspartate aminotransferase abolishes its interactions with mitochondria in vitro. 402 99

The complete amino acid sequence of the mitochondrial aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1) from human heart has been determined based mainly on analysis of peptides obtained by digestion with trypsin and by chemical cleavage with cyanogen bromide. Comparison of the sequence with those of the isotopic isoenzymes from pig, rat and chicken showed 27, 29 and 55 differences, respectively, out of a total of 401 amino acid residues. Evidence for structural microheterogeneity at position 317 has also been obtained.
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PMID:The primary structure of mitochondrial aspartate aminotransferase from human heart. 405 35

The cytoplasmic isozyme of aspartate transaminase is inactivated by trypsin due to loss of a 19-residue peptide from the NH2-terminal region. A second peptide bond at Arg-25 is then cleaved by trypsin leaving a residual core protein, transaminase 26-412. Inactivation by trypsin resembles that for the mitochondrial enzyme (Sandmeier, E., and Christen, P. (1980) J. Biol. Chem. 255, 10284-10289), yet occurs 10 times faster for the cytoplasmic isozyme. In the mitochondrial enzyme, trypsin cleavage produces equal concentrations of proteins missing the first 26 and 31 amino acids. Sequence variation in the NH2-terminal regions can explain such differences. Specifically, the mitochondrial NH2 terminus has no trypsin-susceptible residue at position 19 and is stabilized by an electrostatic interaction between Asp-15 and Arg-292, whereas position 15 is a valyl residue in the cytoplasmic enzyme. Calorimetric data reveal both a decreased transition temperature (Td) and enthalpy (delta Hd) of denaturation in transaminases 20-412 and 26-412. Interaction of substrates with the active site chromophore and differential scanning calorimetry (DSC) reveal that catalytically inactive transaminases 20-412 and 26-412 can bind amino acid substrates and produce spectroscopically detectable conversion of the pyridoxal to the pyridoxamine form of the protein. By contrast, substrate analogs only form enzymatic Michaelis-type complexes.
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PMID:Selective tryptic cleavage of native cytoplasmic aspartate transaminase holoenzyme. 636 6

The influence of a raw green gram (RGG) diet, an autoclaved green gram (AGG) diet and green gram trypsin inhibitors (GGTI) incorporated in AGG diet on urinary and blood urea and creatinine levels in rats was studied. The activities of certain liver enzymes of pathways associated with protein or amino acid metabolism were also studied. The levels of urea and creatinine in urine and blood were found to be significantly increased in rats fed the RGG and GGTI-incorporated AGG diets when compared to the animals fed with the AGG diet. The levels of enzyme activities of arginase, ornithine transcarbamoylase, aspartate aminotransferase and alanine aminotransferase were also found to be significantly increased along with that of urea and creatinine, The possible role of GGTI on the altered levels of the above-mentioned parameters is discussed.
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PMID:Influence of dietary raw green gram (Phaseolus aureus Roxb) and green gram trypsin inhibitors on the activity of certain protein metabolism enzymes in rats. 733 25

Native mitochondrial aspartate aminotransferase (AATase) is cleaved selectively by trypsin at the peptide bonds after Arg 26 or after Lys 31 yielding two shortened enzyme derivatives, AATase 27-410, and AATase 32-410. Recent x-ray crystallographic determination of the spatial structure of AATase has shown that the NH2-terminal segments of the two polypeptide chains of this dimeric enzyme pass in front of the active site clefts and form two separate junctions with the neighboring subunit which are not contiguous with the main subunit interface (Eichele, G., Ford, G. C., Glor, M., Jansonius, J. N., Mavrides, C., and Christen, P. (1979) J. Mol. Biol. 133, 161-180). The peptide bonds cleaved by trypsin are situated in the following stretch of the polypeptide chain which runs in exposed position on the surface of the subunit. The split-off peptide is lost during gel filtration. The molecular activity of AATase 27/32-410 (a mixture of about equal amounts of the two not readily separable derivatives) is about 3% of that of the native enzyme. In contrast, the K'm values for aspartate and 2-oxoglutarate are unchanged, indicating an unaltered geometry of the substrate binding site. A substantially diminished syncatalytic response of the reactivity of Cys 166 toward 5,5'-dithiobis-(2-nitrobenzoate) suggests that the decrease in catalytic activity is due to an interference with the syncatalytic conformational dynamics observed previously in AATase (Gehring, H., and Christen, P. (1978) J. Biol. Chem. 253, 3158-3163). Consonant with a role of the NH2-terminal segment in propagating the syncatalytic conformational rearrangements the rate of the tryptic cleavage is retarded 4-fold in the presence of the transaminating substrate pair aspartate and oxalacetate.
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PMID:Mitochondrial aspartate aminotransferase 27/32-410. Partially active enzyme derivative produced by limited proteolytic cleavage of native enzyme. 743 Jan 25

Amino-acid sequence of kynureninase purified from rat liver cytosol was determined by an amino-acid sequencer. The enzyme was degraded to small peptides with cyanogen bromide, TPCK-trypsin, endoproteinase Glu-C, lysyl endoprotease and alpha-chymotrypsin. The enzyme subunit consisted of 464 amino acids, and the molecular weight of subunit was determined to be 52,510. The coenzyme pyridoxal phosphate-binding residue was lysine of which position was 276, and the N-terminal residue was N-acetylmethionine. The homology search between this enzyme and the other pyridoxal phosphate-dependent enzymes showed that kynureninase was similar to mitochondrial aspartate aminotransferase, and also to cystathionine gamma-synthase and gamma-lyase to a lesser extent.
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PMID:Amino-acid sequence of rat liver kynureninase. 757 21

Various reports have described that amino acid substitutions can alter substrate, positional, inhibitory, and target gene specificities of proteins. By using the method of Chou and Fasman, the present work predicts that critical amino acids for converting these substrate specificities of trypsin, L-lactate dehydrogenase, aspartate aminotransferase, beta-lactamase, and cytochrome P-450 are found to exist within regions predicted as beta-turns. The ratios of hydroxylation and oxygenation positions of substrates by cytochrome P-450 and lipoxygenase, respectively, are varied by changes of the protein structures, probably around turn conformations. Inhibitory specificities of bovine pancreatic trypsin inhibitor and alpha 1-antitrypsin and target gene specificity of glucocorticoid receptor are converted by changing turn structures. Occurrence of beta-turn probabilities can be predicted around the amino acid alteration positions of an evolutionally antecedent protein of a nylon degradation enzyme. These findings will have relevance to work on protein engineering and enzyme evolution.
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PMID:Critical amino acids responsible for converting specificities of proteins and for enhancing enzyme evolution are located around beta-turn potentials: data-based prediction. 813 29


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