Enzymes conjugation with reduced

Cytochrome P-450 and cysteine conjugate p-lyse are primarily localized in the proximal tubules, and these enzymes also contribute to the susceptibility of the proximal tubule to toxicant injury. Specifically, widely used industrial solvents such as chloroform produce tubular nephrotoxicity via cytochrome P-450 activation, and haloaUcanes and haloalkenes (e.g. trichloroethylene) are rendered toxic by cysteine conjugate (3-lyse activation [24,24a]. In addition, overdoses of acetaminophen (APAP) cause nephrotoxicity that is characterized by proximal tubular necrosis [25]. APAP undergoes cytochrome P-450-mediated activation to produce a toxic electrophile, N-acetyl-p-benzoquinon-eimine (NAPQI) [25a]. Although NAPQI is extremely reactive, it is detoxified by conjugation with reduced GSH unless NAPQI is formed in excess of the cellular capacity for GSH conjugation. The excess NAPQI is available to bind to critical cellular proteins and to induce oxidative stress, resulting in disruption of cellular homeostasis and tubular injury [26]. 

Lipoic acid exists as a mixture of two structures a closed-ring disulfide form and an open-chain reduced form (Figure 18.33). Oxidation-reduction cycles interconvert these two species. As is the case for biotin, lipoic acid does not often occur free in nature, but rather is covalently attached in amide linkage with lysine residues on enzymes. The enzyme that catalyzes the formation of the lipoamide nk.2Lg c requires ATP and produces lipoamide-enzyme conjugates, AMP, and pyrophosphate as products of the reaction. 

A final consideration is to provide adequate desalting of the reduced antibody molecule from excess reducing agent. If even a small amount of a thiol-containing reductant remains, subsequent conjugation with a maleimide-activated enzyme will be inhibited. 

Figure 27.14 An oligonucleotide modified with cystamine and reduced to generate a free sulfhydryl may be conjugated with an SPDP-modified enzyme, forming a disulfide linkage.

Metaboiism - Oxcarbazepine is rapidly reduced by cytosolic enzymes in the liver to its MHD, which is primarily responsible for the pharmacological effect of oxcarbazepine. MHD is metabolized further by conjugation with glucuronic acid. 

As has been stated before, oxirane derivatives are formed as intermediates during metabolic oxidations at carbon-carbon double bonds. These epoxides (arene oxides) undergo spontaneous isomerization to phenols, or enzymic hydration via epoxide hydrase to trans- dihydrodiols, or reaction with reduced glutathione (GSH) via specific GSH-transferases to the corresponding conjugates (Scheme 11), which eventually appear in urine... 

The use of capillaries to run immunoassay reactions was presented in the 1980s and 1990s by Halsall group [8] for the detection of atrazine. The system was based on a flow injection analysis protocol where different solutions (sample, conjugate) were injected and reacted in the capillary itself the affinity-captured enzyme conjugate then reacted with a substrate solution injected in the capillary prior to detection. The reacted substrate solution was then pushed towards an electrode for amperometric detection. The authors showed that the incubation time reached equilibrium after about 20 min, but they worked with a capillary of 360 pm diameter. In order to really reduce the incubation time below lmin, we developed disposable polymer chips with an internal diameter of 40 pm (almost one order of magnitude smaller than... 

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