Naslov
Characterization of glutathione-S-transferases in zebrafish (Danio rerio)

Autori
Mihaljević, Ivan ; Glisić, Branka ; Popović, Marta ; Žaja, Roko ; Lončar, Jovica ; Fent, Karl ; Kovačević Radmila ; Smital, Tvrtko

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Molecules of Life - FEBS3+ Meeting / Kos, Janko ; Poklar Ulrih, Nataša - Ljubljana : Slovenian Biochemical Society, 2015, 180-180

ISBN
978-961-93879-0-0

Skup
Molecules of Life - FEBS3+ Meeting

Mjesto i datum
Portorož, Slovenija, 16.09.2015. - 19.09.2015

Vrsta sudjelovanja
Predavanje

Vrsta recenzije
Nije recenziran

Ključne riječi
glutathion-S-transferases ; zebrafish ; characterization

Sažetak
In order to be metabolized and excreted form the organism, xenobiotics need to pass through four phases of the cell metabolism. Detoxification process starts with the phase 0 - initial uptake of xeno- and endobiotics across the cell membrane by polyspecific membrane transporters. After the entrance, the xenobiotics pass through phase I of cell metabolism, which is characterized with enzymatic bioactivation of initial compounds through oxidation-reduction reactions. The phase II encompasses enzymes that mediate conjugation of phase I metabolites or initial compounds to the water soluble moieties, thus increasing the bioavailability of metabolized compounds. Finally, phase III refers to the efflux of parent compounds or metabolites by membrane transporters. Glutathione-S- transferases (GSTs) are one of the key enzymes that mediate phase II of cellular detoxification. The goal of our research was a comprehensive characterization of GSTs in zebrafish (Danio rerio) as a valuable vertebrate model species frequently used in various fields of biology and biomedicine. A detailed phylogenetic analysis of GST superfamily revealed 27 zebrafish gst genes: seven classes of cytosolic Gsts, mitochondrial genes within the kappa class, and microsomal genes within the MAPEG class. Further insights into the orthology relationships between human and zebrafish GSTs/Gsts were obtained by the conserved synteny analysis. Expression of gst genes in six tissues (liver, kidney, gills, intestine, brain and gonads) of adult male and female zebrafish was determined using qRT-PCR. The results showed ubiquitously high expression of gstp, gstm (except in liver), gstr1, mgst3a and mgst3b, high expression of gsto2 in gills and ovaries, gsta in intestine and testes, gstt1a in liver, and gstz1 in liver, kidney and brain. Functional characterization was performed on 9 representatives of cytosolic Gst classes after overexpression in E. coli and subsequent protein purification. Enzyme kinetics was measured using GSH as co-substrate and a series of model substrates. The functional characterization revealed specific interactions with model GST substrates and enabled the comparison of kinetic properties with human and fish orthologs. Finally, we conclude that Gstp1, Gstp2, Gstt1a, Gstz1, Gstr1, Mgst3a and Mgst3b have important role in the biotransformation of xenobiotics, while Gst Alpha, Mu, Pi, Zeta and Rho classes are involved in the crucial physiological processes.

Izvorni jezik
Engleski

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