Protein methylases in Trypanosoma brucei brucei: activities and response to DL-alpha-difluoromethylornithine.
Protein methylases I, II and III were detected in extracts of Trypanosoma brucei brucei, and characterized according to the specific amino substituent methylated. Only protein methylase II activity was elevated by difluoromethylornithine treatment of T. b. brucei, and hence this enzyme was characterized further. Protein methylase II transferred methyl groups from S-adenosyl-L-methionine (S-AdoMet) to the carboxyl residues of several protein substrates, exhibiting highest activity with histone VIII-S (arginine-rich subgroup f3). The crude enzyme had an apparent Km for histone VIII-S of 28 mg ml-1 (11.4 mM-aspartyl and 18.4 mM-glutamyl residues methylated), and an apparent Km for S-AdoMet of 8.4 microM. T. b. brucei protein methylase II was sensitive to inhibition by S-adenosyl-L-homocysteine and its analogue sinefungin with apparent Ki values of 12.9 and 1.6 microM, respectively. Using a partially purified preparation, analysis of kinetic data in the presence and absence of sinefungin indicated that this analogue acts as a competitive inhibitor of the S-AdoMet binding site, and as a non-competitive inhibitor of the (protein) histone VIII-S binding site. The possible role of the enzyme in morphological control and its potential as a chemotherapeutic target are discussed.
Yarlett
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N
Haskins Laboratories, Department of Biology, Pace University, New York, NY 10038.
Quamina
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A
Bacchi
C J
CJ
D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D000518:Eflornithine / Q000494:pharmacology*; D011495:Histone-Lysine N-Methyltransferase / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008745:Methylation; D011497:Protein O-Methyltransferase / Q000378:metabolism*; D011484:Protein-Arginine N-Methyltransferases / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity; D014346:Trypanosoma brucei brucei / Q000187:drug effects / Q000201:enzymology*; D014353:Trypanosomiasis, African / Q000469:parasitology
D011497:Protein O-Methyltransferase; D011484:Protein-Arginine N-Methyltransferases; D011495:Histone-Lysine N-Methyltransferase; D000241:Adenosine; C006235:sinefungin; D000518:Eflornithine