Velocity allosteric effects

Lack of perfect specificity in carrier-solute recognition provides for the possibility that structurally similar solutes may compete for carrier availability. Analysis of competitive [Eq. (18)] and noncompetitive [Eq. (19)] inhibition as well as cooperativity effects (allosteric modulation by structurally dissimilar solutes) on carrier-mediated solute flux is equivalent to assessment of the velocity of enzyme reactions. 

Substrates can affect the conformation of the other active sites. So can other molecules. Effector molecules other than the substrate can bind to specific effector sites (different from the substrate-binding site) and shift the original T-R equilibrium (see Fig. 8-9). An effector that binds preferentially to the T state decreases the already low concentration of the R state and makes it even more difficult for the substrate to bind. These effectors decrease the velocity of the overall reaction and are referred to as allosteric inhibitors. An example is the effect of ATP or citrate on the activity of phosphofructokinase. Effectors that bind specif-... 

For example, Bachelard used [Mgtotai]/[ATPtotai ] = 1 in his rate studies, and he obtained a slightly sigmoidal plot of initial velocity versus substrate ATP concentration. This culminated in the erroneous proposal that brain hexokinase was allosterically activated by magnesium ions and by magnesium ion-adenosine triphosphate complex. Purich and Fromm demonstrated that failure to achieve adequate experimental control over the free magnesium ion concentration can wreak havoc on the examination of enzyme kinetic behavior. Indeed, these investigators were able to account fully for the effects obtained in the previous hexokinase study. ... 

Figure 16.18. Activation of Phosphofructokinase by Fructose 2,6-Bisphosphate. (A) The sigmoidal dependence of velocity on substrate concentration becomes hyperbolic in the presence of 1 lM fructose 2,6-bisphosphate. (B) ATP, acting as a substrate, initially stimulates the reaction. As the concentration of ATP increases, it acts as an allosteric inhibitor. The inhibitory effect of ATP is reversed by fructose 2,6-bisphosphate. [After E. Van Schaftingen, M.F. Jett, L. Hue, and H. G. Hers. Proc. Natl. Acad. Sci. 78(1981) 3483.]...

Figure 4-51 shows the effect of different interaction factors on the velocity curve of an allosteric tetramer. As the interaction factors decrease (i.e., as the cooperativity increases), the curves become more sigmoidal and [S]o.s decreases. 

Relationship between the initial velocity (v) and the substrate concentration [S] for an allosteric enzyme that shows a homotropic effect. The substrate functions as a positive modulator. The profile is sigmoidal, and during the steep part of the profile, small changes in [S] can cause large changes in v. Ko.i represents the substrate concentration corresponding to half-maximal velocity. 

Fig. 20.13. Allosteric regulation of isocitrate dehydrogenase (ICDH). Isocitrate dehydrogenase has eight subunits, and two active sites. Isocitrate, NAD, and NADH bind in the active site ADP and Ca are activators and bind to separate allosteric sites. A. A graph of velocity versus isocitrate concentration shows positive cooperativity (sigmoid curve) in the absence of ADP. The allosteric activator ADP changes the curve into one closer to a rectangular h5 perbola, and decreases the (S0.5) for isocitrate. B. The allosteric activation by ADP is not an all-or-nothing response. The extent of activation by ADP depends on its concentration. C. Increases in the concentration of product, NADH, decrease the velocity of the enzyme through effects on the allosteric activation.

Reactions of this type show limited sensitivity to changes in the concentrations of their substrates and it is often found that the flow of material through the reaction increases despite a decrease in substrate concentration and vice versa. This results from the effect of allosteric regulators in altering the affinity of the enzyme for its substrate and sometimes also the maximum velocity of the reaction. 

Figure 8.7. (a) Simulation of the effects of varying the effective number of active sites in an enzyme (w) on the shape of the initial velocity versus substrate concentration curve for a cooperative enzyme, (b) Simulation of the effects of varying the allosteric constant (L) on the shape of the initial velocity versus substrate concentration curve for a cooperative enzyme. 

FIGURE 6.11 Effect of substrate concentration on the initial velocity of an allosteric enzyme in the presence and absence of specific modulators... 

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