Kinetics of S-adenosylmethionine cellular transport and protein methylation in Trypanosoma brucei brucei and Trypanosoma brucei rhodesiense

African trypanosomes of the Trypanosoma brucei group are agents of disease in man and animals. They present unique biochemical characteristics such as the need for preformed purines and have extensive salvage mechanisms for n ucleoside recovery. In this regard we have shown that trypanosomes have a d edicated transporter for S-adenosylmethionine (AdoMet), a key metabolite in transmethylation reactions and polyamine synthesis. In this study we compa red the apparent kinetics of AdoMet transport, cytosolic AdoMet pool format ion, and utilization of AdoMet in protein methylation reactions using two i solates: Trypanosoma brucei brucei, a veterinary parasite, and Trypanosoma brucei rhodesiense, a human pathogen that is highly refractory and has grea tly reduced susceptibility to standard trypanocidal agents active against T . b. brucei. The apparent K-m, values for [methyl-H-3]AdoMet transport, der ived by Hanes-Woolf analysis, for T. b. brucei was 4.2 and 10 mM for T. b. rhodesiense, and the V-max values were 124 and 400 mu mol/liter/min, respec tively. Both strains formed substantial cytosolic pools of AdoMet, 1600 nmo l/10(9) T. b. brucei and 3500 nmol/10(9) T. b. rhodesiense after 10 min inc ubation with 25 mM exogenous AdoMet. Data obtained from washed trichloroace tic acid precipitates of cells incubated with [methyl-H-3]AdoMet indicated that the rate of protein methylation in T. b. brucei was four-fold greater than in T. b. rhodesiense. These results demonstrate that the unique rapid uptake and utilization of AdoMet by African trypanosomes is an important co nsideration in the design and development of new agents of potential use in chemotherapy. (C) 1999 Academic Press.