Biochemistry: The Chemical Reactions of Living Cells, Volume 2 - D. Metzler 1980

Enzymes: Protein Catalysts of Cells
Fundamentals of Enzyme Kinetics

To measure The rate of a chemical reaction, one must first initiate the reaction at a specific time (e.g., by rapidly mixing the reactants) and then, at strictly controlled Temperature and pH values (which is critical), measure the concentration of a reactant or product after a given time interval (or intervals). Researchers have shown remarkable ingenuity by developing a vast array of Methods to record kinetic curves for individual Enzymes. Regardless of the method chosen by the experimenter, the primary interest lies in the rate at which the concentration of a substance changes over time. Very often, a researcher plots a kinetic curve representing the time dependence of the concentration of a reactant (substrate) [S] or product [P] (Fig. 6-1).

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FIG. 6-1. Kinetic curve for an enzymatic reaction in which substrate S is converted into products. I — tangent line whose slope corresponds to the initial reaction rate; II — tangent to the curve at a point corresponding to a non-zero time; the slope of this line is less than the slope of the tangent drawn at t = 0.

The rate of an enzymatic reaction v is defined as follows:

The rate of a chemical reaction is usually expressed in mol∙(L∙s)^-1 (or M∙s^-1), whereas enzymology more frequently employs the unit mol∙(L∙min)^-12). The instantaneous rate, which is typically of primary interest to the researcher, is determined by the slope of the tangent to the kinetic curve at a given point (Fig. 6-1).

The initial reaction rate is the rate at time zero. In certain cases (for example, with the curve shown in Fig. 6-1), determining the initial rate with sufficient accuracy can be quite challenging.

For A number of Chemical Reactions, such as first-order processes, the plot of the kinetic curve on a semi-logarithmic scale (i.e., the dependence of lg[S] on t) yields a straight line, thus eliminating the need to determine the slope of the tangent at time zero. However, in most cases, activity measurements are designed to yield an approximately linear kinetic curve (at least over short time intervals). This often necessitates The Use of highly sensitive Methods for determining product concentration. Consequently, radioactive substrates are widely used in such experiments.

¹⁾ It should be noted that the equality between the rate of substrate consumption and the rate of product accumulation holds true only under steady-state conditions (Sec. A,5).

²⁾ To express the reaction rate, intensive parameters should always be used (e.g., M∙s^-1) rather than extensive ones (e.g., μmol∙s^-1 without specifying the solution volume).

Note that when the kinetic curve is not linear from the outset, and when The amount of substance reacted over a given time interval is taken as a measure of the rate, inaccurate estimations may result. Sometimes, the integrated form of the rate equation, which describes product accumulation over time, is employed. Other approaches for comparing relative rates also exist, and these are applicable even to the kinetic curve shown in Fig. 6-1.