Aspartate aminotransferase. The effects of ionic concentration on kinetic constants of both isoenzymes

Abstract

1. The Michaelis constants for both isoenzymes for both substrates depend strongly on ionic concentration, being approximately proportional to phosphate concentration over considerable ranges. This is probably an effect of anions only. 2. In the absence of added salt, K(m) (2-oxoglutarate) (anionic isoenzyme) is so small as to be indeterminate. 3. K(m) (l-aspartate) (anionic isoenzyme) passes through a sharp minimum at about 3.3mm-phosphate. It is not clear whether this is a specific effect of phosphate. 4. Both substrates are inhibitory at sufficiently low ionic concentrations. 5. A modified graphical procedure is described for the derivation of the kinetic constants.

The open/closed conformational equilibrium of aspartate aminotransferase

“…In contrast, Tobler et al (1987) had shown that aspartate binds to the bimane-labeled pyridoxamine enzyme (Kd = 440 pM). The discrepancy may be explained by the fact that in the presence of Hepes buffer, aspartate binds more tightly than in the presence of the phosphate buffer used in the present study (see also Experimental Procedures) due to the competing effect of Pi (Boyde, 1968).…”

Hydrogen Exchange at the β‐Carbon of Amino Acids during Transamination

“…With the transaminases, the further rate of amino group transfer back and forth then depends on the binding affinity of the 2-0x0 acid for the enzyme and its concentration when the equilibrium of Eqn (1) is reached. Variable Michaelis constants for the substrates of glutamic-oxaloacetic transaminase have been described [21]. Therefore the values cited in Table 2 are somewhat selective.…”

Ligand Substitution Chemistry and Enzymology

“…This follows from the nature of the ligand substitution reaction equilibrium: The direct proportionality between the surrogate dissociation constant and the concentration of another surrogate is a most useful diagnostic criterion. Moreover, when observed (39,41,44), it offers a direct refutation that appreciable amounts of free or solvated enzyme can be present in the enzymatic reaction.…”