Pure noncompetitive inhibition

Pure noncompetitive inhibition occurs if Ki = Ki. This situation is relatively uncommon the Lineweaver-Burk plot for such an instance is given in Eigure 14.15. Note that K is unchanged by I (the x-intercept remains the same, with or without I). Note also that Tmax decreases. A similar pattern is seen if the amount of enzyme in the experiment is decreased. Thus, it is as if I lowered [E],... 

Pure noncompetitive inhibition is said to exist if i is unaffected by the concentration of substrate. 

This equation corresponds to that for mixed inhibition (Table 9.1) if Kt 4= K r However, if = K t, then pure noncompetitive inhibition is the result. Thus, it can be seen that, mechanistically speaking, pure noncompetitive inhibition is a special case of mixed inhibition. 

The double reciprocal form of Eq. (9.41) for pure noncompetitive inhibition is ... 

This equation predicts that both the slope and the l/v0 intercept of a Lineweaver-Burk plot will increase with increasing inhibitor concentration, but the intercept on the 1/[S]0 axis (-1 Km) will not change. A series of plots for several experiments with different concentrations of inhibitor will all pass through the l/[S]o intercept as shown in Fig. 9-4(6), indicating that pure noncompetitive inhibition does not alter Km. 

Fig. 9-4 Lineweaver-Burk plot Uv0 versus 1/[S]0 for ( ) pure competi,ive inhibition and (b) pure noncompetitive inhibition.

To derive the Lineweaver-Burk equations, we proceed by simply taking the reciprocals of each side of the equations expressing v0 as a function of [S]0 in Table 9-1. The corresponding graphs of l/u0 versus l/[S]o have varying slopes, intercepts, or both as [I] is varied. Pure noncompetitive inhibition shows lines... 

In noncompetitive inhibition (Figure 8.38). the inhibitor can combine with either the enzyme or the enzyme-substrate complex. In pure noncompetitive inhibition, the values of the dissociation constants of the inhibitor and enzyme and of the inhibitor and enzyme-substrate complex are equal (Section 8.5.1). The value of decreased to a new value... 

Depending on the inhibitor being considered, the values of these dissociation constants may or may not be equivalent. There are two forms of noncompetitive inhibition pure and mixed. In pure noncompetitive inhibition, a rare phenomenon, both Kz values are equivalent. Mixed noncompetitive inhibition is typically more complicated because the Kj values are different. 

The mechanism of inhibition is defined by the relative values of Ais and Ay, which are respectively the inhibition constants at [ATP] Km and >> Km.2S 29 Inhibition constants are measures of potency, because they equal the free inhibitor concentration when the rate is reduced by 50%. The mechanism is competitive if inhibition tends to zero when ATP is saturating dATP] Km). This mechanism is seen if Ais Ay. Conversely, the mechanism is uncompetitive if inhibition tends to zero when [ATP] Km, because As Ay. Inhibition is noncompetitive when it occurs both at [ATP] Km and [ATP] Am. Pure noncompetitive inhibition (Ais = Ay) arises when potency is independent of ATP-concentration. Mixed noncompetitive inhibition (Als A Ay) occurs if there is a tendency towards competitive or uncompetitive. 

Figure 4.6 Apparent inhibition constant as a function of ATP-concentration relative to Km for competitive inhibition (solid line), pure noncompetitive inhibition (dashed line) and uncompetitive inhibition (dotted line). The inhibition constant is 5 nM for each mechanism.

In pure noncompetitive inhibition, the inhibitor binds with equal affinity to the free enzyme and to the enzyme-substrate (ES) complex. In noncompetitive inhibition, the enzyme-inhibitor-substrate complex IES does not react to give product P. A kinetic scheme for noncompetitive inhibition is given in Figure 6.41... 

In noncompetitive inhibition (Figure 8.38), the inhibitor can combine with either the enzyme or the enzyme-substrate complex. In pure noncompetitive inhibition, the values of the dissociation constants of the inhibitor and enzyme and of the inhibitor and enzyme—substrate complex are equal (Section 8.5.1). The value of is decreased to a new value called V( i. and so the intercept on the vertical axis is increased. The new slope, which is equal to Km/ V( i. is larger by the same factor. In contrast with Vjjxix. is not affected by pure noncompetitive inhibition. The maximal velocity in the presence of a pure noncompetitive inhibitor, V ax. is given by... 

Lineweaver-Burk plots provide a good illustration of competitive inhibition and pure noncompetitive inhibition (Fig. 9.18). In competitive inhibition, plots of 1/v vs 1/[S] at a series of inhibitor concentrations intersect on the ordinate. Thus, at infinite substrate concentration, or 1/[S] = 0, there is no effect of the inhibitor. In pure noncompetitive inhibition, the inhibitor decreases the velocity even when [S] has been extrapolated to an infinite concentration. However, if the inhibitor has no effect on the binding of the substrate, the is the same for every concentration of inhibitor, and the lines intersect on the abcissa. 

Fig. 9.18. Lineweaver-Burk plots of competitive and pure noncompetitive inhibition. A. lAi versus 1/[S] in the presence of a competitive inhibitor. The competitive inhibitor alters the intersection on the abscissa. The new intersection is 1/K , p (also called 1/K ). A compietitive inhibitor does not affect B. 1/Vj versus 1/[S] in the presence of a pure noncompetitive inhibitor. The noncompetitive inhibitor alters the intersection on the ordinate, Wmax.app W niax> But docs not offcct 1/K j. A pure noncompetitive inhibitor binds to E and ES with the same affinity. If the inhibitor has different affinities for E and ES, the lines will intersect above or below the abscissa, and the noncompetitive inhibitor will change both the and the V, .

To derive the Lineweaver-Burk equations, we proceed by taking the reciprocals of each side of Eqs. (5.25), (5.29), and (5.32). The corresponding graphs of 1/vq versus l/[S]o have various characteristic changes in slopes and intercepts as [I] is varied. Competitive inhibition gives lines that all intersect on the ordinate. Pure noncompetitive inhibition (in which K, = K,) gives lines that all intersect on the abscissa. For anti- or uncompetitive inhibition, the telltale feature is that the set of lines are all parallel to each other. For mixed inhibition [K K in Eq. (5.32)], both the slopes and the intercepts on the ordinate and abscissa differ for different values of [I] see Fig. 5-31. 

Pure noncompetitive inhibition (decrease in V ax with no change in K ) is seldom observed in enzyme kinetics studies, except in the case of very small inhibitors, such as protons, metal ions, and small anions. For noncompetitive... 

Consider the pure noncompetitive inhibition by two nonexclusive inhibitors, depicted by Eq. (5.35). If there are two dead-end inhibitors I and X, the kinetic question is whether an EIX complex forms, and if so, whether the dissociation constant of I from EIX is the same as from El (and similarly for X from EIX and EX). To answer this, the substrate concentration is held constant, the concentration of the two inhibitors is varied, and initial velocities are determined (Yonetani Theorell, 1965 Yonetani, 1982). 

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