Mechanisms of inactivation

In order to clear up the mechanism of inactivation of excited states, we examined the processes of quenching of fluorescence and phosphorescence in PCSs by the additives of the donor and acceptor type253,2S5,2S6 Within the concentration range of 1 x 1CT4 — 1 x 10"3 mol/1, a linear relationship between the efficiency of fluorescence quenching [(/0//) — 1] and the quencher concentration was found. For the determination of quenching constants, the Stem-Volmer equation was used, viz. 

The prominent role played by Japanese investigators in carbohydrate science is underscored by the two substantial chapters by Japanese authors in the current volume. This volume also pays tribute to one of the greatest Japanese carbohydrate scientists, Hamao Umezawa, in the obituary article contributed by Tsuchiya, Maeda, and Horton. Hamao Umezawa dedicated his entire, extraordinarily productive career to the development of antibiotics his innovative contributions are exemplified by his chapter in Volume 30 of this series on the biochemical mechanism of inactivation of aminoglycoside antibiotics. 

Villar-Palasi C On the mechanism of inactivation of muscle glycogen phosphorylase by insulin. Biochim Biophys Acta 1994 1224 384. 

Bloomfield S.F. Arthur M. (1994) Mechanisms of inactivation and resistance of spores to chemical... 

SCHEME 11.2 Postulated mechanism of inactivation of a serine protease by a functionalized dihydrocoumarin such as molecule 3 (i i = benzyl).28... 

Because of the multiple degradation pathways that may take place at elevated temperature, protein stability monitoring data may not conform to the Arrhenius relationship, and the maximum temperature selected for accelerated stability studies must be carefully selected. Gu et al. [32] described the different mechanisms of inactivation of interleukin-1 (3 (IL-1 (3) in solution above and below 39°C. In this example, the multiple mechanisms precluded the prediction of formulation shelf life from accelerated temperature data. In contrast, by working at 40° C and lower, Perlman and Nguyen [33] were able to successfully extrapolate data from stability studies of tissue plasminogen activator down to 5°C. 

The shaking of protein solutions may lead to aggregation and precipitation as a result of several mechanisms, such as air oxidation, denaturation at the interface, adsorption to the vessel, or mechanical stress. These possibilities were systematically examined for solutions of human fibroblast interferon [50]. In this example, mechanical stress was identified as the causative factor in the inactivation. The proposed mechanism of inactivation by mechanical stress was through orientation of the asymmetrical protein in the... 

Figure 83 Plot of fcobs as a function of inactivator concentration for a single-step mechanism of inactivation.

Hence, for any irreversible enzyme inactivator, we can quantify the effectiveness of inactivation using the second-order rate constant kanJKi. This constant thus becomes the key metric that the medicinal chemist can use in exploring the SAR of enzyme inactivation by a series of compounds. In terms of individual rate constants, the definitions of both nact and A) depend on the details of the mechanisms of inactivation, as will be described below. 

Our second example of drugs that function as mechanism-based inactivators is the steroid 5a-reductase inhibitors finasteride and dutasteride. The mechanism of inactivation by these compounds is an interesting departure from the typical target enzyme covalent modification seen with most mechanism-based inactivators. 

Because duration of activity of the catecholamines is significantly longer than that of neuronally released norepinephrine, the effects on tissues are more prolonged. This difference has to do with the mechanism of inactivation of these substances. Norepinephrine is immediately removed from the neuroeffector synapse by way of reuptake into the postganglionic neuron. This rapid removal limits duration of the effect of this neurotransmitter. In... 

Parker D.J., and Allison, W.S. (1969) The mechanism of inactivation of glyceraldehyde 3-phosphate dehydrogenase by tetrathionate, o-iodosobenzoate, and iodine monochloride. J. Biol. Chem. 244, 180-189. 

C2. Carson, P. E., Schrier, S. L., and Kellermeyer, R. W., Mechanism of inactivation of glucose-6-phosphate dehydrogenase in human erythrocytes. Nature 184, 1292-1293 (1959). 

Baxa, U., Speransky, V., Steven, A. C., and Wickner, R. B. (2002). Mechanism of inactivation on prion conversion of the Saccharomyces cerevisiae Ure2 protein. Proc. Natl. Acad. Sci. USA 99, 5253-5260. 

Guo Z, Wagner CR, Hanna PE. 2004. Mass spectrometric investigation of the mechanism of inactivation of hamster arylamine N-acetyitransferase 1 by N-hydroxy-2-acetylami-nofluorene. Chem Res Toxicol 17 275. 

Mivacurium chloride (Mivacron) is a newer agent that is chemically related to atracurium. The primary mechanism of inactivation is hydrolysis by plasma cholinesterase. Although it is useful for patients with renal or hepatic disease, some caution is warranted, since these individuals may have reduced plasma cholinesterase as a result of the disease. Mivacurium has an onset of action (1.8 minutes) and duration of effect (20 minutes) only twice that of succinylcholine, and in this respect, it is the most similar to succinylcholine of all of the nondepolarizing agents. 

Silverman and coworkers have carried out extensive research on the mechanism of inactivation of MAO by cyclopropylamine analogues. They first reported in the early 1980s that A/-cyclopropyl-A/-arylalkylamines are mechanism-based inactivators of MAO [121,125], The mechanism proposed was enzyme-catalyzed one electron oxidation of A/-cyclopropylamines to give reactive ring-opened products which further react with either flavin and/or a cysteine residue, depending on the structure of the inactivator. According to their reports [120,125, 126], 1-phenylcyclopropylamine (50) attached reversibly to a cysteine residue and irreversibly to the flavin when it activated MAO B, whereas 50 modified only the flavin during inactivation of MAO A. In the case of frans-2-phenylcyclopropy-lamine (8a) and A/-cyclopropyl-a-methylbenzylamine (51), both MAO A and B are inactivated by attachment to a cysteine residue (Fig. 3). 

Kennedy RD, Quinn IE, Johnston PG et al. BRCA1 mechanisms of inactivation and implications for management of patients. Lancet 2002 360 1007-1014. 

Shvartsman, P.Y. Sharygina, N.V. (1982) Study of the mechanism of inactivation and muta-... 

The reaction of sulfur-containing biomolecules with platinum antitumor compounds, thereby preventing binding to the critical DNA target, is a possible mechanism of inactivation and is supported by numerous studies. Thus, glutathione (GSH, a cysteine-containing tripeptide see also Fig. 6), which is the predominant intracellular thiol and is present in concentrations varying from 0.5 to 10 mM, is able to inhibit the reaction of DNA with [Pt(en)Cl2] (74) and with cis-Pt (75, 76). It has also been observed that the presence of cysteine can inhibit the reaction between cis-Pt and d-Guo (77). Furthermore, the antitumor activity of cis-Pt was proved to be inhibited by coadministered methionine (78, 79) and even a bis-adduct between cis-Pt and methionine has been isolated from the urine of patients (80). 

A second mechanism of inactivation might be the reaction of sulfur-containing biomolecules with the cis-Pt-DNA monoadducts (product 1 in Fig. 4), which prevents those from rearranging to toxic bifunctional adducts. Supportive for such a mechanism is the observation that GSH can be cross-linked to DNA by cis-Pt (41,41a) and [Pt(en)Cl2] (74), and that cysteine can be cross-linked to d-Guo by cis-Pt (77). Furthermore, cis-Pt-DNA monoadducts can be experimentally quenched with thiourea, which reduces drug toxicity (82, 83). trans-Pt also yields monofunctional adducts after reaction with DNA, and these rearrange somewhat slower than does cis-Pt into bifunctional adducts (41,84), clearly for sterical reasons. The relatively long-living monofunctional adducts react efficiently with GSH and proteins (41 a, 84-86). 

Detailed kinetic studies to determine the rate-limiting step in the binding of Pt to S. This will be important in understanding the mechanisms of inactivation and resistance in normal tissues and tumors. 

Fig. 5. Proposed mechanism of inactivation of a-chymotrypsin by the pseudoirreversible inhibitor 2-ethoxy-4.H-3,l-benz-oxazin-4-one. A substituted...

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