Structural features of cytochrome P450 1A associated with the absence of EROD activity in liver of the loricariid catfish Pterygoplichthys sp |
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Authors: | Parente Thiago E M Rebelo Mauro F da-Silva Manuela L Woodin Bruce R Goldstone Jared V Bisch Paulo M Paumgartten Francisco J R Stegeman John J |
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Institution: | a Lab. Biologia Molecular Ambiental, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Brasilb Lab. de Física Biológica, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Brasilc Biology Department, Woods Hole Oceanographic Institution, USAd Lab. de Toxicologia Ambiental, Escola Nacional de Saúde Pública, Fundação Oswaldo Cruz, Brasil |
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Abstract: | The Amazon catfish genus Pterygoplichthys (Loricariidae, Siluriformes) is closely related to the loricariid genus Hypostomus, in which at least two species lack detectable ethoxyresorufin-O-deethylase (EROD) activity, typically catalyzed by cytochrome P450 1 (CYP1) enzymes. Pterygoplichthys sp. liver microsomes also lacked EROD, as well as activity with other substituted resorufins, but aryl hydrocarbon receptor agonists induced hepatic CYP1A mRNA and protein suggesting structural/functional differences in Pterygoplichthys CYP1s from those in other vertebrates. Comparing the sequences of CYP1As of Pterygoplichthys sp. and of two phylogenetically related siluriform species that do catalyze EROD (Ancistrus sp., Loricariidae and Corydoras sp., Callichthyidae) showed that these three proteins share amino acids at 17 positions that are not shared by any fish in a set of 24 other species. Pterygoplichthys and Ancistrus (the loricariids) have an additional 22 amino acid substitutions in common that are not shared by Corydoras or by other fish species. Pterygoplichthys has six exclusive amino acid substitutions. Molecular docking and dynamics simulations indicate that Pterygoplichthys CYP1A has a weak affinity for ER, which binds infrequently in a productive orientation, and in a less stable conformation than in CYP1As of species that catalyze EROD. ER also binds with the carbonyl moiety proximal to the heme iron. Pterygoplichthys CYP1A has amino acid substitutions that reduce the frequency of correctly oriented ER in the AS preventing the detection of EROD activity. The results indicate that loricariid CYP1As may have a peculiar substrate selectivity that differs from CYP1As of most vertebrate. |
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Keywords: | AHR Aryl hydrocarbon receptor BNF ß -naphthoflavone CYP1A Cytochrome P450 1A DMBA 7 12-dimethylbenz[α]anthracene DMSO Dimethyl sulfoxide ER Ethoxyresorufin EROD Ethoxyresorufin-O-deethylase i p intraperitoneally ITS internal transcribed spacer MD Molecular dynamics NADPH Reduced Nicotinamide adenine dinucleotide phosphate OD Optical density PCB126 3 3&rsquo 4 4&rsquo 5-pentachlorinated biphenyl PCR Polymerase Chain Reaction PDB Protein data bank PMDB Protein Model Data Bank qPCR Quantitative real time PCR RACE reactions Rapid ampli?cation of cDNA ends RMS Root mean square RMSD Root-mean-squared deviation RMSF Root-mean-squared fluctuation SRS Substrate recognition site TRIS tris(hydroxymethyl)aminomethane VTFM Variable target function method |
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