Feedback inhibition enzyme kinetics

Aiming at a computer-based description of cellular metabolism, we briefly summarize some characteristic rate equations associated with competitive and allosteric regulation. Starting with irreversible Michaelis Menten kinetics, the most common types of feedback inhibition are depicted in Fig. 9. Allowing all possible associations between the enzyme and the inhibitor shown in Fig. 9, the total enzyme concentration Er can be expressed as... 

Although the MM equation is a powerful kinetic form to which the vast majority of enzyme kinetics has been fitted, one should not forget the assumptions and limitations of the model. As a basic example, feedback inhibition, whereby the product of the reaction inhibits the enzyme-substrate cooperativity, multiple-substrate reactions, allosteric modifications, and other deviations from the reaction scheme in equation (1) are treated only adequately by the MM formalism under certain experimental conditions. In other words, enzyme kinetics are often bent to conform to the MM formalism for the sake of obtaining a set of parameters easily recognizable by most biochemists. The expUcit mathematical and experimental treatment of reaction mechanisms more complex than that shown in equation (1) is highly involved, although a mathematical automated kinetic equation derivation framework for an arbitrary mechanism has been described in the past (e.g., ref. 6). 

To fully establish whether arogenate (41), phenylpyruvate (39), or both were pathway intermediates to Phe (1) in vascular plants, it was essential to unambiguously identify all enzymatic processes (and encoding genes) needed for conversion of prephenate (38) into Phe (1) both in vivo and in vitro. Moreover, it was also essential to obtain rigorous enzymatic kinetic data using highly purified recombinant enzymes in vitro for all potential substrates, in order to compare and contrast relative efficacies/feedback inhibition properties and so forth. 

Enzyme production is governed by a cell s genes. Enzyme activity is further controlled by pH changes, alterations in the concentrations of essential cofactors, feedback inhibition by the products of the reaction, and activation by another enzyme, either from a less active form or an inactive precursor ( zymogen). Such changes may themselves be under the control of hormones or the nervous system. See also enzyme kinetics. 

It is a monomeric protein of M.W. about 70,000, shows Kj, values for L-tryptophan and dimethylallyl pyrophosphate of 0.067 and 0.2 mM, respectively, and seems to have a relatively low turnover number, about 7 sec . During studies on this enzyme it was observed (13) that agroclavlne and elymoclavine, the terminal alkaloids in the strain used for the isolation of the enzyme, inhibited purified DMAT synthetase. At concentrations of 3 mM ( v<750 mg/1) agroclavlne and elymoclavine inhibited the enzyme 90% and 70%, respectively. The inhibition is of a mixed or uncompetitive type as shown by kinetic analysis with either tryptophan or dimethylallyl pyrophosphate as the variable substrate (Fig. 6). Subsequently, feedback Inhibition by elymoclavine was also demonstrated by GrSger s group (3) for chanoclavlne cyclase and by us for anthranllate synthetase from... 

Thymidine kinase serves as a salvage reaction in the phosphorylation of thymidine to yield dTMP. Its activity is under allosteric control as revealed by Okazaki and Kornberg with a highly purified preparation from E. coli [186]. Sigmoidal kinetics are obtained with ATP as substrate, and this is converted to a hyperbolic form by dCDP which functions as an activator. The affinity for thymidine is also increased by dCDP. Feedback inhibition is obtained with dTTP, the end product of the pathway. Inhibition by dTTP is competitive with the phosphate acceptor, thymidine, and noncompetitive with the donor, ATP. The kinetics of inhibition in the presence of the activator are difficult to interpret in terms of a simple competition between dTTP and dCDP for an allosteric site. This type of control apparently serves the same function as that described for dCMP deaminase above where the activity of the enzyme is decreased by the end product, dTTP, and increased when other deoxyribonucleotides accumulate. 

Purified preparations of thymidine kinase have also been obtained from animal tumors [187,188]. These are similar to the E. coli enzyme in their susceptibility to feedback inhibition by dTTP, but no activators other than ATP have been identified. Sigmoidal kinetics with ATP as substrate are obtained with an aggregated form of the enzyme, but not with the disaggregated enzyme. Both forms of the enzyme show complex inhibition by dTTP. Thymidine kinase has also been studied in crude extracts of Tetrahymena pyriformis [189], and in this case no inhibition by dTTP was evident. 

Hundreds of metabohc reac tions take place simultaneously in cells. There are branched and parallel pathways, and a single biochemical may participate in sever distinct reactions. Through mass action, concentration changes caused by one reac tion may effect the kinetics and equilibrium concentrations of another. In order to prevent accumulation of too much of a biochemical, the product or an intermediate in the pathway may slow the production of an enzyme or may inhibit the ac tivation of enzymes regulating the pathway. This is termed feedback control and is shown in Fig. 24-1. More complicated examples are known where two biochemicals ac t in concert to inhibit an enzyme. As accumulation of excessive amounts of a certain biochemical may be the key to economic success, creating mutant cultures with defective metabolic controls has great value to the produc tion of a given produc t. 

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