Protein microsomal

Ruangyuttikam, W. Skiles, G. L. Yost, G. S. Identification of a cysteinyl adduct of oxidized 3-methylindole from goat lung and human liver microsomal proteins. Chem. Res. Toxicol. 1992, 5, 713-719. 

Fig. 4 HLM Clint, free vs clogD. HLM Clint, app corrected for microsomal protein binding using a computational model for microsomal binding. Open squares and filled triangles represent the same chemical series as in Fig. 3 (series A and B, respectively)...

Blonder, J., Rodriguez-Galan, M.C., Chan, K.C., Lucas, D.A., Yu, L.R., Conrads, T.P, Issaq, H. J., Young, H.A., Veenstra, T.D. (2004). Analysis of murine natural killer cell microsomal proteins using two-dimensional liquid chromatography coupled to tandem electrospray ionization mass spectrometry. J. Proteome Res. 3, 862-870. 

Figure 2. Bioavailability of soot-adsorbed benzo(a)pyrene (BaP). The transfer of l4C-BaP from diesel soot to microsomes was measured. Microsomal protein (0.5 mg) was incubated at 37 C with the [14C]-benzo(a)pyrene-coated diesel particles. ( ) lung microsomes and 0.2 mM NADPH (O) lung microsomes (A) liver microsomes with 0.2 mM NADPH (A) liver microsomes ( ) 0.5 mg albumin ( ) buffer (0.15 M phosphate buffer, pH 7.7, containing 3 mM MgCI2 and 0.1 mM EDTA). The presence of NADPH, a cofactor necessary for BaP metabolism, did not affect the transfer of BaP from the soot. (Reproduced with permission from ref. 15. Copyright 1988 Elsevier.)...

Although metal-catalyzed protein oxidation is undoubtedly a very effective oxidative process, the origin of free metal ions under in vivo conditions is still uncertain (see Chapter 21). However, protein oxidation can probably be initiated by metal-containing enzymes. Mukhopadhyay and Chatterjee [31] have shown that NADPH-stimulated oxidation of microsomal proteins was mediated by cytochrome P-450 and occurred in the absence of free metal ions. It is important that in contrast to metal ion-stimulated oxidation of proteins, ascorbate inhibited and not enhanced P-450-dependent protein oxidation reacting with the oxygenated P-450 complex. The following mechanism of P-450-dependent oxidation of the side chain protein amino acid residues has been proposed ... 

Choi et al. [32] showed that NADPH stimulated the modification of microsomal proteins, which was probably dependent on lipid peroxidation and inhibited by iron chelators. 

On the other hand, in rats, a single dose of 6,156 mg/kg hexachloroethane in mineral oil had no effects on a different set of biochemical indicators of liver function (microsomal protein, oxidative demethylase, NADP-NT reductase, glucose-6-phosphatase, or lipid conjugated diene concentration) when measured 2 hours after compound administration (Reynolds 1972). Each of these parameters is an indicator of microsomal function. The authors postulated that the observed lack of effects could have been the result of slow uptake of hexachloroethane by the liver in a 2-hour period. Gastrointestinal absorption of hexachloroethane in mineral oil is probably minimal because, unlike olive oil, mineral oil cannot be digested. Dissolved lipophilic materials could be excreted in the feces soon after administration because mineral oil can act as a laxative. Thus, the author s hypothesis that minimal hexachloroethane would reach the liver in 2 hours is reasonable. 

D.K. Han, J. Eng, H. Zhou and R. Aebersold, Quantitative profiling of differentiation-induced microsomal proteins using isotope-coded affinity tags (ICAT) and mass spectrometry, Nat. Biotech., 19 (2001) 946-951. 

Tran, T.H., Von Moltke, L.L., Venkatakrishnan, K., Granda, B.W., Gibbs, M.A., Obach, R.S., Harmatz, J.S. and Greenblatt, D.J. (2002) Microsomal protein concentration modifies the apparent inhibitory potency of CYP3A inhibitors. Drug Metabolism and Disposition, 30 (12), 1441-1445. 

Pittz EP, Abraham R, Rourke D, et al. 1978. Effect of oral administration to mice of 30 ppm of mirex on the sodium dodecyl sulfate polyacrylamide gel electrophorectic patterns of hepatic microsomal proteins. Toxicol Appl Pharmacol 45(1) 335-336. 

Pittz EP, Rourke D, Abraham R, et al. 1979. Alterations in hepatic microsomal proteins of mice administered mirex orally. Bull Environ Contam Toxicol 21 344-351. 

Figure 3. Effect of temperature on conversion of molinate to its sulfoxide by carp liver microsomal mfo (liver microsomal protein 6 mg/mL, pH 7.4, NADPH 3 mg incubated for 15 min)...

Figure 4. Effect of carp liver microsomal protein concentration on its mfo activity (NADPH 3 mg/mL, pH 7.4 incubated for 15 min at 25°C)...

Figure 5. Effect of incubation time on the carp liver microsomal mfo activity (NADPH 2 mg/mL, pH 7.4 incubated at 25° C. Microsomal protein concentration...

Table I). The levels of both, cytochrome P-L50 (Table i) and its NADPH (reduced nicotinamide adenine dinucleotide phosphate) requiring reducing component (Figure l)(which can be measured as NADPH dependent cytochrome c reductase) are substantial in fish liver microsomes, although lower than in mammals. NADPH cytochrome c reductase level in trout Salmo trutta lacustris) is 20 nmol cytochrome c reduced/mg microsomal protein/min the corresponding activity in male Sprague Dawley rat liver microsomes is 96 nmol cytochrome c reduced/mg microsomal protein/min (lU). 

Several properties of hepatic microsomal AHH activity were compared in control and DBA-pretreated male little skates as shown in Table I. Following treatment there was an approximately 35-fold increase in specific enzyme activity, as quantitated by fluorescence of the phenolic metabolites formed (3, 21). The pH optimum, which was fairly broad, and the concentration of benzo(a)-pyrene (0.06 mM) that had to be added to the incubation mixture to achieve maximum enzyme activity were the same for both control and induced skate hepatic microsomes. The shorter periods observed for linearity of product formation with microsomes from the induced skates is thought to be related to the much higher AHH activity present, and may be due to substrate depletion or the formation of products which are inhibitory (i.e., compete with the MFO system as they are substrates themselves). A similar explanation may be relevant for the loss of linear product formation at lower microsomal protein concentrations in the induced animals. 

Arylhydrocarbon (benzo[a]pyrene) hydroxylase, benzphetamine-N-demethylation, ethylmorphine-N-demethylation, ethoxycoumarin-0-deethylation and ethoxyresorufin-0-deethylation were performed by published procedures (31,32,33,34), but optimized for use with trout microsomes as described previously (30, 35). Hemoprotein P-450 was determined by the procedure of Estabrook et al. (36) to avoid spectral interference by hemoglobin. Microsomal protein content was estimated either by the method of Ross and Shatz (37) or Lowry et al. (38), using bovine serum albumin standards. 

Each pellet ( dirty microsomal fraction ) was resuspended in 2 mL of ice-cold 1.15 % KCl, collected together and ultracentrifuged again at 100 000 x g for 45 min at 0 °C. The supernatant was discarded and the pellet was rinsed thrice with ice-cold dilution buffer. The rinsed pellet was resuspended in ice-cold 1.15 % KCl to yield a final concentration 20 mg mL of microsomal proteins. The microsomal suspension was immediately aliquoted, frozen and stored at —80 °C. 

Addya S, Anandatheerthavarada HK, Biswas G, Bhagwat SV, MuUick J, et al. 1997. Targeting of NH2-terminal-processed microsomal protein to mitochondria a novel pathway for the biogenesis of hepatic mitochondrial P450MT2. J Cell Biol 139 589-599. 

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