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Query: UMLS:C0027960 (mole)
 21,279 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The binding of CTP and ATP to aspartate transcarbamylase at pH 7.8 and 8.5 at 25 degrees has been investigated by equilibrium dialysis and flow microcalorimetry. The binding isotherms for CTP at both pH 7.8 and 8.5 and ATP AT PH 8.5 can be fit by a model which assumes three tight, three moderately tight, and six weak binding sites. The binding isotherms for ATP at pH 7.8 are best fit by a model which assumes six tight and six weaker sites. Both finite differenceH binding and finite differenceS binding are negative for both nucleotides at both pH values, so that the binding is enthalpy driven. For both nucleotides, finite differenceH is the same for the first two classes of binding sites, implying that the difference in the dissociation constants of these two classes of sites is the result of entropic effects. Direct pH measurements and calorimetric measurements in two buffers with very different heats of ionization (Tris and Hepes) indicate that the binding of both nucleotides is accompanied by the binding of protons. In the pH range 6.7-8.4, the number of moles of protons bound per mole of nucleotide increases as the pH decreases.
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PMID:Calorimetric analysis of aspartate transcarbamylase from Escherichia coli: binding of cytosine 5'-triphosphate and adenosine 5'-triphosphate. 23 71

THE COOPERATIVITY AND FEEDBACK INHIBITION EXHIBITED BY THE REGULATORY ENZYME, ASPARTATE TRANSCARBAMOYLASE (CARBAMOYLPHOSPHATE: L-aspartate carbamoyltransferase; EC 2.1.3.2), from Escherichia coli are generally attributed to ligand-promoted conformational changes involving alterations in the subunit interactions. However, no quantitative estimates have been made of the effect of ligands on the strength of the intersubunit bonding domains. The native enzyme, composed of two catalytic trimers "crosslinked" by three regulatory dimers, shows little tendency to dissociate in neutral buffers at room temperature. In addition, very little exchange was observed in 2 hr between subunits within the intact enzyme and free subunits. Although exchange was enhanced in solutions of low ionic strength containing the bisubstrate analog, N-(phosphonacetyl)-L-aspartate, the rates of exchange were too small to permit reliable estimates of the weakening of the bonding domains caused by the ligand. Studies were conducted, therefore, on a less stable oligomeric complex which resembles the native enzyme in structure and allosteric behavior but lacks one regulatory subunit. These molecules, containing only four bonding domains between the catalytic and regulatory polypeptide chains (compared to six in the native enzyme), disproportionate to form the more stable native enzyme and free catalytic subunits. An electrophoretic technique is described for measuring the rate of disproportionation which is controlled by the rupture of the intersubunit bonding domains. This rate is enhanced about 300-fold upon the addition of the active-site ligand. Hence the ligand-promoted allosteric conversion of the enzyme-like complex from the constrained to the relaxed conformation involves a substantial weakening of the intersubunit interactions corresponding to about 1.7 kcal/mole (7.1 kJ/mole) per bonding domain between a catalytic and a regulatory chain.
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PMID:Ligand-promoted weakening of intersubunit bonding domains in aspartate transcarbamolylase. 33 46

The interaction between the catalytic subunit (c3) and the regulatory subunit (r2) of aspartate transcarbamylase from Escherichia coli was studied by measuring the reversible formation of the c3r6 complex as a function of r2 concentration. Conversion to the native enzyme was prevented by using a very low concentration of c2 (40 ng per ml) in the presence of bovine serum albumin. A simple hyperbolic r2 saturation curve was obtained suggesting the presence of only one kind of c:r domain. From the association constant for the formation of c3r6, the free energy of c:r interaction can be estimated to be about -10 Cal per mole. Neither CTP nor ATP appears to affect the strength of c:r interaction in this complex. Succinate in the presence of carbamyl phosphate promotes tighter binding. At higher concentration of c3 and nonsaturating levels of r2, conversion to the native enzyme (c3r6) takes place. This renaturation process is second order with respect to the concentration of c3 and is virtually irreversible. Renaturation is inhibited by saturating levels of r2 and to some extent by both CTP and ATP. The effect of ligands on c:r interactions reported here may have significance in the allosteric mechanism of the native enzyme.
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PMID:Subunit interactions in aspartate transcarbamylase. The interaction between catalytic and regulatory subunits and the effect of ligands. 108 46