EVALUATION OF OPTIMUM CONDITIONS OF 2-SUBSTRATE ENZYME-REACTIONS

  • 1 January 1977
    • journal article
    • research article
    • Vol. 15  (8) , 405-410
Abstract
A modern approach was described for the evaluation of the optimal conditions for 2-substrate enzyme reactions. It chiefly involved the determination of the substrate concentration for the primary reaction and the catalytic concentration of indicator enzymes. The interrelationships between the concentration of the 2 substrates (concentration pairs) were described mathematically to be hyperbolic, and, in case of competitively inhibited reactions, to be parabolic. Calculated optimum concentrations were rechecked experimentally for the reactions of aspartate aminotransferase and alanine aminotransferase. For pyridine coenzyme linked indicator reactions it could be demonstrated that they mostly follow zero order kinetics. One of the products of the primary reaction reacted, in its steady state concentration, as the 2nd substrate. This represented the size of the lag phase of the coupled reaction. The Km of this substance must be known in order to calculate the catalytic concentration of the indicator enzyme in relation to that of the primary enzyme. Its concentration could be fixed arbitrarily within certain limits, depending on whether the calculated result actually could be realized; otherwise a larger lag phase must be tolerated. For practical reasons, it was generally possible to measure only a certain percentage of maximum reaction rate.

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