Three-dimensional structure at 5 Å resolution of cytosolic aspartate transaminase from chicken heart

Borisov, V.V.; Borisova, S.N.; Качалова, Г.С.; Sosfenov, N. I.; Vaĭnshteĭn, B. K.; Torchinsky, Yu.M.; Braunstein, A. E.

doi:10.1016/0022-2836(78)90403-5

Cited by 45 publications

(11 citation statements)

References 27 publications

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“…The lack of reactivity of the Cys-390 in the apoenzyme subunit of the hybrid molecule confirms the result obtained with a carboxymethylated hybrid [23]. Since crystallographic studies [24,25] have shown that the active sites are 30 A apart, it is not likely that the binding of one coenzyme blocks directly the Cys-390 of the other subunit. More probably, this binding induces a conformational change which is transmitted via subunit interactions to the other active site, so that the Cys-390 cannot react with NbS2.…”

Section: Discussionsupporting

confidence: 77%

A rapid preparation of the apo—holo hybrid of aspartate aminotransferase

Vergé

Arrio-Dupont

1981

FEBS Letters

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Section: Discussionsupporting

confidence: 77%

A rapid preparation of the apo—holo hybrid of aspartate aminotransferase

Vergé

Arrio-Dupont

1981

FEBS Letters

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“…Recently, we have shown that mAATase(pig) and mAATase(chicken) have identical chain folds and, in a 3.2-A resolution difference map between the two structures, we have observed strong local electron density differences that could be associated with substitutions of individual amino acid side chains (36). The structures of cAATase(chicken) and mAATase(chicken) as seen at low resolution are closely related (20,23). The dimers have the same overall shape and the helices on their surfaces form similar patterns.…”

Section: H H H H H H H H H H H H H H H H H H H H H H H H H H H mentioning

confidence: 68%

Three-dimensional structure of a pyridoxal-phosphate-dependent enzyme, mitochondrial aspartate aminotransferase.

Ford¹,

Eichele²,

Jansonius³

1980

Proc. Natl. Acad. Sci. U.S.A.

245

139

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X-ray diffraction studies to 2.8A resolution have yielded the three-dimensional structure of mitochondrial aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1), an isologous a2 dimer (M, = 2 X 45,000) The subunits are rich in secondary structure and contain two domains, one of which anchors the coenzyme, pyridoxal 5'-phosphate. Each active site lies between the subunits and is composed of residues from both of them.Aspartate aminotransferase (AATase; L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1) is the most studied of the vitamin B-6-dependent enzymes (1, 2). These enzymes catalyze a wide variety of transformations in amino acid metabolism. AATase effects the reversible transfer of an amino group from L-aspartate or L-glutamate to the a-keto acids a-ketoglutarate and oxalacetate. In the course of the double displacement reaction (3, 4) the coenzyme shuttles between the pyridoxal-P form (bound via an aldimine linkage to the E-amino group of a lysine) and the pyridoxamine-P form. "Syncatalytic" conformational changes occur in the enzyme matrix (5-7). Various coenzyme-substrate intermediates are identified by characteristic absorption and circular dichroism spectra (2, 8). The stereochemistry of pyridoxyl-catalyzed reactions has been probed (9), and a dynamic reaction mecha'nism has been proposed (10).Two homologous, genetically independent isozymes of AATase have been found in animal tissues, one in the cytosol (cAATase), the other in the mitochondrial matrix (mAATase). Both are a2 dimers of about 2 X 400 amino acids. The amino acid sequences of the pig (11-13) and chicken (refs. 14 and 15; U. Hausner, K. J. Wilson, and P. Christen, personal communication) isozymes are known or have been almost completely elucidated. Crystallized AATase offers an exceptional opportunity for the study of interactions amongst protein, coenzyme, and substrates during catalysis. The crystalline enzyme is catalytically competent (16). Single-crystal microspectrophotometric studies on cAATase (17, 18) and mAATase (16,19) have permitted the recognition of several reaction intermediates and the evaluation of some dissociation constants and kinetic parameters (19). Now a detailed picture of the enzyme is emerging from X-ray studies of such crystals. The high-resolution X-ray analyses of cAATase from chicken (20) and pig (21) are near completion. Here we report the three-dimensional structure of chicken mAATase in the internal aldimine form (pyridoxal-5'-P-enzyme) as revealed by a 2.8-A resolution X-ray study. RESULTS Structure determinationChicken mAATase crystallizes in a triclinic unit cell that contains one a2 dimer (22) whose subunits are related by a noncrystallographic dyad (23). Diffraction data to 2.8-A resolution of the native protein and the derivatives listed in Table 1 were collected on a CAD4F diffractometer (Enraf-Nonius, Delft, Netherlands). Previously found heavy atom sites (23) were reevaluated by difference Fourier maps. They were refined by alternate cycles of multiple i...

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“…This selective permeability may be the basis for the unique intracellular localisation of the isozymes [3] and work is in progress to account for the selectivity in molecular terms; it has already been shown that modification of a single exposed sulphydryl group abolishes uptake of the mitochondrial isozyme into mitochondria [lo]. Further light will no doubt be shed on this and other comparative aspects of aspartate aminotransferases by work currently in progress on the crystal structures of the cytosolic isozymes from pig [ l l ] and chicken [12] and of the mitochondrial isozyme from chicken [13,14], and on the primary structures of the isozymes from a variety of sources (chicken mitochondrial [15], turkey mitochondrial [ 161, human mitochondrial and cytosolic [17], bovine mitochondrial [18] and the single enzyme from Eschericlziu coli [19j).…”

mentioning

confidence: 99%

The Cytosolic and Mitochondrial Aspartate Aminotransferases from Pig Heart. A Comparison of their Primary Structures, Predicted Secondary Structures and Some Physical Properties

Barra¹,

Bossa²,

Martini³

et al. 1980

Eur J Biochem

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1.A primary structure is presented for mitochondrial aspartate aminotransferase from pig heart based on previously published results [Barra et al. (1977) FEBS Lett. 83, 241 -244; ( 1 979 3. Hydrophobic chromatography of the isozymes using alkyl agaroses suggests that the cytosolic but not the mitochondrial isozyme has a hydrophobic patch or pocket capable of binding to the alkyl side chains of the affinity medium.4. Evidence is presented to show that the cytosolic isozyme contains some basic amino acids whose ionizations are not expressed under native conditions. The mitochondrial isozyme does not show this effect.5. Predictions of the secondary structures of the isozymes are presented based on four predictive models. Minimum predictions are derived on the basis of agreement of three out of four of the individual predictions. These agregate predictions underestimate the amount of ordered secondary structure in both isozymes and suggest a relatively low degree of similarity between the isozymes. Individual predictive models, on the other hand, reveal greater similarities in secondary structures between the two isozymes.Aspartate aminotransferase, in common with a small number of other enzymes, exists in two distinct molecular forms one associated with the cell cytosol and the other with the mitochondria [l-31. These two isozymes differ in chemical and physical properties (for a review, see [4]) and indeed it is now known that, in humans, they are coded by genes on different chromosomes (I. Craig, quoted in [5]). Some time ago we [6] and another group [7] reported the complete primary structure of the cytosolic isozyme from pig heart and it seemed logical to extend these investigations to the corresponding mitochondrial form in order to provide a basis for understanding the comparative enzymology, evolutionary history and compartmen-

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Three-dimensional structure at 5 Å resolution of cytosolic aspartate transaminase from chicken heart

Cited by 45 publications

References 27 publications

A rapid preparation of the apo—holo hybrid of aspartate aminotransferase

A rapid preparation of the apo—holo hybrid of aspartate aminotransferase

Three-dimensional structure of a pyridoxal-phosphate-dependent enzyme, mitochondrial aspartate aminotransferase.

The Cytosolic and Mitochondrial Aspartate Aminotransferases from Pig Heart. A Comparison of their Primary Structures, Predicted Secondary Structures and Some Physical Properties

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