Inhibition protein modification

Pyridoxamine is a potent inhibitor of the irreversible rearrangement of the initial product of reversible glycation to the advanced glycation end-product. Such inhibitors of this reaction are collectively known as amadorins, and the name pyridorin has been coined for pyridoxamine used in this way (KhaUfah et al., 1999). Pyridoxamine also inhibits protein modification caused by lipid peroxides in both cases, it seems to act by trapping carbonyl compounds formed as intermediates (Onorato et al., 2000 Voziyan et al., 2002). 

The role of the Arg at the - 2 or - 3 position is unclear and may be presequence-dependent as studies on a variety of precursors revealed that mutating the Arg results in cleavage inhibition or modification in some cases, but not in others. It may be that the structure, rather than the primary sequence composition of the presequence and perhaps of the mature protein, determines the MPP cleavage site (Gakh et al. 2002). 

The preceding experiments prove that there is an intermediate on the reaction pathway in each case, the measured rate constants for the formation and decay of the intermediate are at least as high as the value of kcat for the hydrolysis of the ester in the steady state. They do not, however, prove what the intermediate is. The evidence for covalent modification of Ser-195 of the enzyme stems from the early experiments on the irreversible inhibition of the enzyme by organo-phosphates such as diisopropyl fluorophosphate the inhibited protein was subjected to partial hydrolysis, and the peptide containing the phosphate ester was isolated and shown to be esterified on Ser-195.1516 The ultimate characterization of acylenzymes has come from x-ray diffraction studies of nonspecific acylenzymes at low pH, where they are stable (e.g., indolylacryloyl-chymotrypsin),17 and of specific acylenzymes at subzero temperatures and at low pH.18 When stable solutions of acylenzymes are restored to conditions under which they are unstable, they are found to react at the required rate. These experiments thus prove that the acylenzyme does occur on the reaction pathway. They do not rule out, however, the possibility that there are further intermediates. For example, they do not rule out an initial acylation on His-57 followed by rapid intramolecular transfer. Evidence concerning this and any other hypothetical intermediates must come from additional kinetic experiments and examination of the crystal structure of the enzyme. 

Chloramphenicol and secobarbital exhibit properties similar to those of tienilic acid, but they have not been studied in humans (11). Oxidative dechlorination of chloramphenicol with formation of reactive acyl chlorides appears to be an important metabolic pathway for irreversible inhibition of CYP. Chloramphenicol binds to CYP, and subsequent substrate hydroxylation and product release are not impaired. The inhibition of CYP oxidation and the inhibition of endogenous NADPH oxidase activity suggest that some modification of the CYP has taken place, which inhibits its ability to accept electrons from the CYP reductase (11). Secobarbital completely inactivates rat CYP2B1 functionally, with partial loss of the heme chromophore. Isolation of the N-alkylated secobarbital heme adduct and the modified CYP2B1 protein revealed that the metabolite partitioned between heme N-alkylation, CYP2B1 protein modification, and epoxidation. A small fraction of the prosthetic heme modifies the protein and contributes to the CYP2B1 inactivation (12). 

Voziyan PA, Metz TO, Baynes JW, and Hudson BG (2002) A post-Amadori inhibitor pyri-doxamine also inhibits chemical modification of proteins by scavenging carbonyl intermediates of carbohydrate and lipid degradation. Journal of Biological Chemistry 277, 3397- 03. 

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