Aspartate aminotransferase sequence

Many enzymes exist within a cell as two or more isoenzymes, enzymes that catalyze the same chemical reaction and have similar substrate specificities. They are not isomers but are distinctly different proteins which are usually encoded by different genes.22 23 An example is provided by aspartate aminotransferase (Fig. 2-6) which occurs in eukaryotes as a pair of cytosolic and mitochondrial isoenzymes with different amino acid sequences and different isoelectric points. Although these isoenzymes share less than 50% sequence identity, their internal structures are nearly identical.24-27 The two isoenzymes, which also share structural homology with that of E. coli,28 may have evolved separately in the cytosol and mitochondria, respectively, from an ancient common precursor. Tire differences between them are concentrated on the external surface and may be important to as yet unknown interactions with other protein molecules. 

Absorption bands at 500 nm. With many PLP enzymes certain substrates and inhibitors cause the appearance of intense and unusually narrow bands at 500 nm. Such a band is observed with aspartate aminotransferases acting on eryf/zro-3-hydroxyaspartate (Fig. 14-9). This substrate undergoes transamination very slowly, and the 500-nm absorbing form which accumulates is probably an intermediate in the normal reaction sequence. A similar spectrum is produced by tryptophan indole-lyase acting on the competitive inhibitor L-alanine. Under the same conditions the... 

Aspartate aminotransferase 57s, 135s, 753 absorption spectra 749 active site structure 744 atomic structure 750 catalytic intermediates, models 752 NMR spectra 149 quinonoid intermediate 750 Ramachandran plot 61 sequence 57 transamination 742 Aspartate ammonia-lyase 685 Aspartate carbamoyltransferase 348s active sites 348 regulation 540... 

Fig. 1.2 Comparison of sequences of aspartate aminotransferases from different species. The residues are numbered according to the sequence of pig cAspAT.13 Possible deletions introduced into the sequences are indicated by hyphens. Residues that are invariant in pig c- and m-AspAT s are boxed, those that are invariant in pig AspAT s and coli AspAT are indicated as bold letters, and those that are directly interacting with substrate or coenzyme are marked as. ...

Aspartate aminotransferase is the prototype of a large family of PLP-dependent enzymes. Comparisons of amino acid sequences as well as several three-dimensional structures reveal that almost all transaminases having roles in amino acid biosynthesis are related to aspartate aminotransferase by divergent evolution. An examination of the aligned amino acid sequences reveals that two residues are completely conserved. These residues are the lysine residue that forms the Schiff base with the pyridoxal phosphate cofactor (lysine 258 in aspartate aminotransferase) and an arginine residue that interacts with the a-carboxylate group of the ketoacid (see Figure 23.11). 

The cytosolic (cAAT) and mitochondrial (mAAT) isozjmes of aspartate aminotransferase share a significant degree of sequence homology... 

Comparison of amino acid sequences suggests that many other PLP-dependent enzymes have folding patterns similar to those of aspartate aminotransferase but that there are four or more additional different folding patterns Among the enzymes resem-... 

In two-substrate enzyme-catalyzed reactions with a double-displacement reaction sequence, high concentrations of the second substrate may compete with the first substrate for binding. For example, in the reaction catalyzed by aspartate aminotransferase,... 

The sequences of the first lineage appear to be more closely related to those of aromatic-amino-acid aminotransferase, tyrosine aminotransferase, and alanine aminotransferase than to the aspartate aminotransferases of the second group. 

The crystal structure of kynureninase from P. fluorescens was solved in 2004. The enzyme shares the same structural fold as aspartate aminotransferase, but shares low sequence similarity. An active site arginine residue (Arg-375) was identified, which is important in substrate binding. The structure of the human kynureninase, which shows a catalytic preference for 3-hydroxy-kynurenine over L-kynurenine, was solved in 2007. The human enzyme shares the same fold as the P. fluorescens enzyme, and also contains an active site arginine residue (Arg-434). The catalytic mechanism requires two acid/base residues, which have not yet been unambiguously assigned. The hydrolytic cleavage step is believed to proceed via a general base mechanism. ... 

Aspartate aminotransferase has been available in a highly purified state, and in abundance from several sources [31] for more than 2 decades. The primary amino acid sequence [32] of the enzyme as well as its tertiary structure at 2.8 A resolution [33] is now known. It is therefore not surprising that our current view on the mechanism of action of pyridoxal-P-dependent reactions in general and aminotransferases in particular is primarily based on the extensive physicochemical studies which have been performed on aspartate aminotransferase [34],... 

Homology sequence modeling between ALAS and enzymes of the a-family of PLP-dependent enzymes indicated that arginine-439 (R439) of murine ALAS2 is a conserved residue in this family of PLP-dependent enzymes (Figure 2-5). Moreover, this conserved arginine in several enzymes, e.g. aspartate aminotransferase, for... 

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