System noncompetitive inhibition

JIP-1 is a JNK pathway scaffold protein essential for JNK activation in some systems. Like many substrates, JIP-1 binds to JNK in the region generally called the common docking site in MAPKs (28). With the identification of the key residues of JNK required for interaction with the kinase interaction motif of JIP-1, small peptide inhibitors (TI-JIP RPKRPTTLNLF) derived from JIP have been described (29). As expected, TI-JIP was found to be competitive with respect to the phosphoaccep-tor substrate c-Jun and to exhibit noncompetitive inhibition with respect to ATP. It is not yet clear whether peptidomimetics or small molecule inhibitors that might have clinical applicability will be developed using information from JNK-JIP-1 interactions. 

This system can be considered a mixture of competitive and noncompetitive inhibition. The reciprocal form of the rate equation for Eqn. 7.38 is given by Eqn. 7.39. 

Competitive or noncompetitive inhibition of uptake or excretory systems. 

Competitive inhibition occurs when two or more different substrates compete for access to the same microbial enzyme system. Both competitive and noncompetitive inhibition may result in the degradation of one substrate being repressed in the presence of another. 

The noncompetitive inhibition of the decomposition of hydrogen peroxide by cyanide is not immediately obvious from the above reaction mechanism for if cyanide can compete in the formation of the peroxide complex which is responsible for the oxygen evolution in step IV, competitive inhibition might be expected. However, under the experimental conditions necessary to observe peroxide decomposition, an excess of peroxide is required and this is sufficient to give the maximal concentration of the peroxide complex, 1.2 or 1.6 moles of bound peroxide for each erythrocyte or bacterial catalase molecule respectively, i.e., the peroxide complex concentration is independent of the peroxide concentration. Analysis of the system under these conditions shows noncompetitive inhibition to hold. 

Even though it is tempting to consider inhibition of allosteric enzymes in the same fashion as nonallosteric enzymes, much of the terminology is not appropriate. Competitive inhibition and noncompetitive inhibition are terms reserved for the enzymes that behave in line with Michaelis-Menten kinetics. With allosteric enzymes, the situation is more complex. In general, two types of enzyme systems exist, called K systems and V systems. A K system is an enzyme for which the substrate concentration that yields one-half is altered by the presence... 

Fig. 7.1 Chemical reaction mechanism representing a biochemical NAND gate. At steady state, the concentration of species 85 is low if and only if the concentrations of both species Ii and I2 are high. All species with asterisks are held constant by buffering. Thus, the system is formally open although there are two conservation constraints. The first constraint conserves the total concentration of S3 -F 84 -F 85, and the second conserves -F 87. All enzyme-catalyzed reactions in this model are governed by simple Michaelis-Menten kinetics. Lines ending in over an enzymatic reaction step indicate that the corresponding enzyme is inhibited (noncom-petitively) by the relevant chemical species. We have set the dissociation constants, Kp j, of each of the enzymes Ei-Eg, from their respective substrates equal to 5 concentration units. The inhibition constants, K i and K 2, for the noncompetitive inhibition of E1 and 7 by 11 and I2, respectively, are both equal to 1 unit. The Vmax for both Ej and E2 is set to 5 units, and that for E3 and E4 is 1 unit/s. The Vmax s for E5 and Eg are 10 and 1 units/s, respectively. (From [1].)...

As a mle, a noncompetitive inhibition occurs only if there are more than one substrate or product (Todhunter, 1979 Fromm, 1995). For example, a noncompetitive inhibition will take place in a random bisubstrate reaction, when an inhibitor competes with one substrate while the other substrate is varied. Thus, the equilibria shown below describe a Rapid Equilibrium Random bisubstrate system in which an inhibitor competes with A but allows B to bind. 

Figure 2. Noncompetitive inhibition. Rapid Equilibrium Random bisubstrate system with an inhibitor noncompetitive with B. Graphical presentation of Eq. (S-io), with A as a constant and B as a variable substrate.

In the general case, when both complexes EAB and EABI are c ytically active, the inhibitor constants are conveniently obtained from replots of i/A Slope and i/A Intercept versus 1/7, obtained at saturating concentration of one constant substrate, while the other is varied. Under sueh conditions, the system behaves as a single substrate, hyperbolic noncompetitive inhibition system. In this case, the inhibition with respeet to a varied ligand cannot be overcome by saturation with the nonvaried ligand. 

An example for the second (Eq. (6.26)) and the third case (Eq. (6.27)), is also described in Chapter 11 (Section 11.4). It occurs in a Steady-State Ordered Bi Bi system with a dead-end EAP-complex. In this system, the inhibition by the product P is S-parabolic /-linear noncompetitive when the substrate B is variable (and A constant), and the rate equation has the form of Eq. (6.27). The inhibition pattern becomes S-linear /-parabolic noncompetitive, when A is variable (and B constant), and the rate equation takes the form of Eq. (6.26). 

When A is varied, the resulting F term appears in the Intercepti/A function, and when B is the varied substrate, the resulting F term appears in the Slopei/A function. According to the nomenclature of Cleland (Chapter 6 Section 6.6), the system can be described as 6 -hnear, /-parabolic (nonintersecting) noncompetitive inhibition when A is the varied substrate and B is unsaturating, changing to /-linear uncompetitive inhibition when B is saturating. When B is varied, we obtain an -parabolic, /-linear noncompetitive pattern (Fig. 9). 

The reaction rate will be slower owing to the removal of enzyme from the system. The El complex will be catalytically inert. The EIS complex may, however, be susceptible to reconversion to ES and make some contribution to catalytic activity. Noncompetitive inhibition cannot be reversed by excess substrate, but it may be reversed by exhaustive dialysis. 

In freshly isolated rat hepatocytes, Na -dependent cysteine transport is not inhibited by MeAIB and is restricted to System ASC (10, 23). However, the reciprocal is not true, that is, cysteine does inhibit System A activity (10). Like glutamine and histidine, cysteine represents a specific test substrate for a Na" -dependent system other than System A, yet it cannot be used as a system-specific inhibitor because of its noncompetitive inhibition of System A. 

The characteristics for the noncompetitive inhibition of System A have not been described in detail. This type of inhibition suggests that the protein may contain an additional amino acid binding site, other than the substrate site. Enzyme studies indicate that the inhibitor can bind to either the free carrier or the carrier-substrate complex to form a carrier-substrate-inhibitor complex which is inactive (37). 

The physiological significance of the noncompetitive inhibition of System A activity is still subject to speculation. One can estimate the effect of glutamine s inhibitory action [K, = 12 mM, see (9)] on the uptake of alanine using values of 0.26 mM, 4 mM, and 2.2 nmol mg" protein 30... 

The nonnucleoside reverse transcriptase inhibitors (NNRTIs), used in the treatment of AIDS, provide interesting examples of clinically relevant noncompetitive inhibitors. The causative agent of AIDS, HIV, belongs to a virus family that relies on an RNA-based genetic system. Replication of the vims requires reverse transcription of the viral genomic RNA into DNA, which is then incorporated into the genome of the infected host cell. Reverse transcription is catalyzed by a virally encoded nucleic acid polymerase, known as reverse transcriptase (RT). This enzyme is critical for viral replication inhibition of HIV RT is therefore an effective mechanism for abrogating infection in patients. 

Following the initial isolation of the Hnl from M. esculenta [33] in which the peptide sequence was established, an overexpressed version of this enzyme (in E. coli) was prepared [41]. This system is not limited for enzyme quantity (as outlined in Sect. 2.3), and can accept a wide range of aromatic, heterocyclic and aliphatic aldehydes, as well as ketones, as substrates. In practical terms, a measure of the degree of enzyme inhibition by substrates is of significant value and for this system this has been quantified for a range of aldehydes, ketones and alcohols [70]. It was deduced that ketones and alcohols are competitive inhibitors, whilst aldehydes are noncompetitive inhibitors. 

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