Characterization of O(6)-methylguanine-DNA-methyltransferase (O(6)-MGMT) activity in Xiphophorus fishes.
We utilized a custom-synthesized double-strand oligonucleotide containing a single O(6)-methylguanine (O(6)-MG) residue within a restriction endonuclease recognition site to determine O(6)-methylguanine-DNA-methyltransferase (O(6)-MGMT) activity in various tissue extracts prepared from Xiphophorus fish. The results suggest Xiphophorus fish O(6)-MGMT activity has many of the same characteristics as Escherichia coli and mammalian O(6)-MGMT's including rapid reaction kinetics consistent with stoichiometric removal of methyl groups, but exhibits a temperature optimum of 23 degrees C. Results from protein extract activity assays indicate O(6)-MGMT activity patterns among four Xiphophorus tissues followed the order: brain> or =testes>gill> or =liver. In mammals, O(6)-MGMT activity is high in liver, while activity in brain is minimal (i.e. approximately 9% of liver); however, we report that in the Xiphophorus fishes examined, brain tissue extracts exhibited much higher (approximately six-fold) O(6)-MGMT activity levels than liver. Comparison of O(6)-MGMT activity between Xiphophorus species employed in tumor induction experiments did not indicate significant differences in ability to clear the pre-mutagenic O(6)-MG from the oligonucleotide substrate.
Walter
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Department of Chemistry and Biochemistry, Southwest Texas State University, 419 Centennial Hall, 601 University Drive, San Marcos, TX 78666, USA. rw12@swt.edu
Sung
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Intano
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Walter
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D000818:Animals; D001483:Base Sequence; D003532:Cyprinodontiformes / Q000378:metabolism*; D018736:DNA Adducts / Q000378:metabolism; D004260:DNA Repair; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology; D005260:Female; D006801:Humans; D007700:Kinetics; D008297:Male; D008322:Mammals; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D013045:Species Specificity; D013379:Substrate Specificity; D013696:Temperature; D014018:Tissue Distribution
D018736:DNA Adducts; D004791:Enzyme Inhibitors; D009838:Oligodeoxyribonucleotides; C088636:O(6)-benzyl-2'-deoxyguanosine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D003849:Deoxyguanosine