Molecular Dynamics Analysis Reveals Structural Insights into Mechanism of Nicotine N-Demethylation Catalyzed by Tobacco Cytochrome P450 Mono-Oxygenase

CYP82E4, a cytochrome P450 monooxygenase, has nicotine N-demethylase (NND) activity, which mediates the bioconversion of nicotine into nornicotine in senescing tobacco leaves. Nornicotine is a precursor of the carcinogen, tobacco-specific nitrosamine. CYP82E3 is an ortholog of CYP82E4 with 95% sequence identity, but it lacks NND activity. A recent site-directed mutagenesis study revealed that a single amino acid substitution, i.e., cysteine to tryptophan at the 330 position in the middle of protein, restores the NND activity of CYP82E3 entirely. However, the same amino acid change caused the loss of the NND activity of CYP82E4. To determine the mechanism of the functional turnover of the two molecules, four 3D structures, i.e., the two molecules and their corresponding cys–trp mutants were modeled. The resulting structures exhibited that the mutation site is far from the active site, which suggests that no direct interaction occurs between the two sites. Simulation studies in different biological scenarios revealed that the mutation introduces a conformation drift with the largest change at the F-G loop. The dynamics trajectories analysis using principal component analysis and covariance analysis suggests that the single amino acid change causes the opening and closing of the transfer channels of the substrates, products, and water by altering the motion of the F-G and B-C loops. The motion of helix I is also correlated with the motion of both the F-G loop and the B-C loop and; the single amino acid mutation resulted in the curvature of helix I. These results suggest that the single amino acid mutation outside the active site region may have indirectly mediated the flexibility of the F-G and B-C loops through helix I, causing a functional turnover of the P450 monooxygenase

Drug Metabolism in Human Brain: High Levels of Cytochrome P4503A43 in Brain and Metabolism of Anti-Anxiety Drug Alprazolam to Its Active Metabolite

Cytochrome P450 (P450) is a super-family of drug metabolizing enzymes. P450 enzymes have dual function; they can metabolize drugs to pharmacologically inactive metabolites facilitating their excretion or biotransform them to pharmacologically active metabolites which may have longer half-life than the parent drug. The variable pharmacological response to psychoactive drugs typically seen in population groups is often not accountable by considering dissimilarities in hepatic metabolism. Metabolism in brain specific nuclei may play a role in pharmacological modulation of drugs acting on the CNS and help explain some of the diverse response to these drugs seen in patient population. P450 enzymes are also present in brain where drug metabolism can take place and modify therapeutic action of drugs at the site of action. We have earlier demonstrated an intrinsic difference in the biotransformation of alprazolam (ALP) in brain and liver, relatively more α-hydroxy alprazolam (α-OHALP) is formed in brain as compared to liver. In the present study we show that recombinant CYP3A43 metabolizes ALP to both α-OHALP and 4-hydroxy alprazolam (4-OHALP) while CYP3A4 metabolizes ALP predominantly to its inactive metabolite, 4-OHALP. The expression of CYP3A43 mRNA in human brain samples correlates with formation of relatively higher levels of α-OH ALP indicating that individuals who express higher levels of CYP3A43 in the brain would generate larger amounts of α-OHALP. Further, the expression of CYP3A43 was relatively higher in brain as compared to liver across different ethnic populations. Since CYP3A enzymes play a prominent role in the metabolism of drugs, the higher expression of CYP3A43 would generate metabolite profile of drugs differentially in human brain and thus impact the pharmacodynamics of psychoactive drugs at the site of action

Meso- and macrozooplankton communities in the Weddell Sea, Antarctica

The present paper describes composition and abundance of meso- and macrozooplankton in the epipelagic zone of the Weddell Sea and gives a systematic review of encountered species regarding results of earlier expeditions. Material was sampled from 6 February to 10 March 1983 from RV Polarstern with a RMT 1+8 m (320 and 4500 μm mesh size). In agreement with topography and water mass distribution three distinct communities were defined, clearly separated by cluster analysis: The Southern Shelf Community has lowest abundances (approx. 9000 ind./1000 m3). Euphausia crystallorophias and Metridia gerlachei are predominating. Compared with the low overall abundance the number of regularly occurring species is high (55) due to many neritic forms. Herbivores and omnivores are dominating (58% and 35%). The North-eastern Shelf Community has highest abundances (about 31 000 ind./1000 m3). It is predominated by copepodites I–III of Calanus propinquus and Calanoides acutus (61%). The faunal composition is characterized by both oceanic and neritic species (64). Fine-filter feeders are prevailing (65%). The Oceanic Community has a mean abundance of approximately 23 000 ind./1000 m3, consisting of 61 species. Dominances are not as pronounced as in the shelf communities. Apart from abundant species like Calanus propinquus, Calanoides acutus, Metridia gerlachei, Oithona spp. and Oncaea spp. many typical inhabitants of the Eastwind Drift are encountered. All feeding types have about the same importance in the Oceanic Community

Structure-function of cytochromes P450 and flavin-containing monooxygenases - Implications for drug metabolism

This article is a report on a symposium held at Experimental Biology '98 in San Francisco, California. Recent developments in site-directed mutagenesis, computer-modeling, and mechanistic analysis of cytochromes P450 and flavin-containing monooxygenases are described. A unifying theme is the elaboration of general approaches for understanding and predicting the function of individual forms of these enzymes. A related goal is the production of soluble forms of mammalian cytochromes P450 for X-ray crystallography