Stable inhibitory complex

Microsomal monooxygenase inhibitors that form stable inhibitory complexes with P450, such as SKF-525A, piperonyl butoxide, and other methylenedioxphenyl compounds, and amphetamine and its derivatives, can be readily investigated in this way. This is because the microsomes isolated from pretreated animals have a reduced capacity to oxidize many xenobiotics. 

Irreversible inhibition, which is much more important toxicologically, can arise from various causes. In most cases the formation of covalent or other stable bonds or the disruption of the enzyme structure is involved. In these cases the effect cannot be readily reversed in vitro by either dialysis or dilution. The formation of stable inhibitory complexes may involve the prior formation of a reactive intermediate that then interacts with the enzyme. An excellent example of this type of inhibition is the effect of the insecticide synergist piperonyl butoxide (Figure 9.6) on hepatic microsomal monooxygenase activity. This methylenedioxyphenyl compound can form a stable inhibitory complex that blocks CO binding to P450 and also prevents substrate oxidation. This complex results from the formation of a reactive intermediate, which is shown by the fact that the type of inhibition changes from competitive to irreversible as metabolism, in the... 

While it might he sjttculated that a similar biphasic ctlcct on metabolism could occur whenever a lipophilic xenobiotic capable of forming a stable inhibitory complex is administered, lew cases have been well documented. Even in the case of MDP interactions, the isoform spec if icily of cither inhibition or induction has not been well defined. Since both detoxification and activation of chemicals are. to a greater or lesser extent, isoform-sped lie, more recent studies have focused on clarifying which iso forms of P450 arc involved and the importance of these isoforms in the biphasic response (Lewandow ski ei a .. 

Little is knowm of the P450 isozyme specificity for either PBO inhibition, metabolite-inhibitory complex formation or type III spectrum formation. Studies involving five isoforms purified from mouse liver (Ijcvi and Hodgson. 19X5 Be nine I rt til., 1985 showed that epoxidation of aldrin by any of these isofonns was inhibited by PBO and that all isoforms appeared to form a stable inhibitory-complex, However, the form of the type III spectrum was variable, with little correlation between the extent of inhibition and the nature of the spectrum. More recently, Pappas and Franklin (1996 have provided indirect evidence that... 

The reaction is proposed to proceed from the anion (9) of A/-aminocatbonylaspattic acid [923-37-5] to dehydrooranate (11) via the tetrahedral activated complex (10), which is a highly charged, unstable sp carbon species. In order to design a stable transition-state analogue, the carboxylic acid in dihydrooronate (hexahydro-2,6-dioxo-4-pyrimidinecarboxylic acid) [6202-10-4] was substituted with boronic acid the result is a competitive inhibitor of dibydroorotase witb a iC value of 5 ]lM. Its inhibitory function is supposedly due to tbe formation of tbe charged, but stable, tetrabedral transition-state intermediate (8) at tbe active site of tbe enzyme. 

The hydroxyl group bonded to a chiral carbon atom in compound 134 represents a possible site for the formation of a stable enzyme-inhibitor complex with adenosine deaminase (ADA), as in 5-9-(2 -hydroxypropyl) adenine and its T-alkyl derivatives. Moreover, the influence of the nitrogen atom on C-3 regarding the inhibitory activity against ADA deserves further investigation. The substituent on C-2 of the nucleus constitutes the third structural element, whose influence on the biological activity of the molecule must also be considered (89JHC39). 

Rifampicin was first shown by Hartmann et al. 54 to have a specific inhibitory effect on RNA polymerase from E. coli. Later, other active ansamycins were found and RNA polymerases from a large variety of bacteria other than E. coli proved to be sensitive to the drug. More recently, an RNA polymerase from E. coli containing only one subunit and probably involved in the initiation of DNA replication (dna G gene product) has been shown to be resistant to rifampicin5 s This holds true also for the various mammalian RNA polymerases. In contrast to non-specific inhibitors of transcription such as actinomycin and mitomycin, rifampicin interacts specifically with the bacterial enzyme itself. With the aid of 14C-labelled rifampicin it could be shown that the drug forms a very stable complex with the enzyme in a molar ratio of 1 1S6> 57 The dissociation constant of this complex is 10-9 M at 37 °C and... 

The most widely known chemotherapeutic agent directed against TS is 5-fluorour-acil (5-FU). 5-FU was first used clinically almost 50 years ago, yet still remains a mainstay for the treatment of carcinoma of the breast and gastrointestinal tract. In cells, 5-FU is metabolized to 5-FdUMP, which forms a stable inhibitory ternary complex with the co-substrate N5N10-methylene-5,6,7,8-tetrahydrofolate (CH2H4-folate) and TS. In this complex, a covalent bond links the thiol of cysteine 195 of human TS to C6 of deoxyuracil monophosphate (dUMP) and the methylene carbon of the co-substrate is joined to C5 of the nucleotide [55]. The fluorine at C5, unlike the proton, cannot... 

A final group of covalent small-molecule inhibitors of proteases are mechanism-based inhibitors. These inhibitors are enzyme-activated irreversible inhibitors, and they involve a two-hif mechanism that completely inhibits the protease. Some isocoumarins and -lactam derivatives have been shown to be mechanistic inhibitors of serine proteases. A classic example is the inhibition of elastase by several cephalosporin derivatives developed at Merck (Fig. 8). The catalytic serine attacks and opens the -lactam ring of the cephalosporin, which through various isomerization steps, allows for a Michael addition to the active site histidine and the formation of a stable enzyme-inhibitor complex (34). These mechanism-based inhibitors require an initial acylation event to take place before the irreversible inhibitory event. In this way, these small molecules have an analogous mechanism of inhibition to the naturally occurring serpins and a-2-macroglobin, which also act as suicide substrates. 

Antibiotics with Antineoplastic Activity - A number of tumor -inhibitory antibiotics form stable complexes with DNA thereby interfering with synthesis of RNA. Three of these antibiotics, actinomycin, mithramycin and daunomycin (which is probably identical to rubidomycin) possess... 

The last column of table 3 shows the values of k as calculated by the above expression from the values of Vmt C and t given in the same table. It is very probable, therefore, that k expresses the specific inhibitory power of anthraquinone in the reactions studied. Furthermore, anthraquinone seems to behave as a true negative catalyst. Since anthraquinone is not destroyed during the oxidation of anethol, how can its effect be explained in the light of the recently proposed chain mechanism of negative catalysis It is well known that quinones are highly reactive substances and are found to combine with several other organic substances to form imstable and frequently stable complexes. In other words,... 

Inhibitor-PIPs block access to the catalytic site and thereby indiscriminately prevent the dephosphorylation of all PPl substrates. They can form a stable dimeric complex with PPl but are also known to form trimeric complexes with PPl and subsets of PIPs. The best characterized inhibitory PIPs are Inhibitor-2, Inhibitor-1, CPT17, and their paralogs. 

---