Levels of the polyamines spermidine and putrescine and the major intracellular thiols glutathione (GSH), glutathionylspermidine (GSH-SPD) and dihydrotrypanothione [bis-(glutathionyl)spermidine); T[SH]2] were measured by high performance liquid chromatography throughout the growth cycle of the insect trypanosomatid Crithidia fasciculata. The amount of total spermidine, putrescine and glutathione (free and conjugated to spermidine) was found to be elevated during growth. Of the total spermidine, 30 to 50% was found conjugated to glutathione during the exponential growth phase, increasing to 60 to 70% at stationary phase. T[SH]2 was the principal intracellular thiol during exponential growth (12·1 to 17·4 nmol per 108 cells), whereas GSH-SPD was the major thiol in stationary phase (26.2 nmol per 108 cells). GSH levels changed little during the growth cycle and represented a constant proportion (10 to 12%) of the total intracellular glutathione. On dilution of stationary phase cells into fresh medium, a rapid decrease in GSH-SPD levels was observed to be associated with synthesis of T[SH]2. This process reached 90% completion by 15 min, with steady state achieved by 120 min. As the total spermidine and glutathione pools did not increase during this interval, it could be calculated that this rapid redistribution of metabolites resulted in the release of 13 nmol per 108 cells unconjugated spermidine without de novo synthesis. This mechanism for rapidly elevating the intracellular concentration of free spermidine may be advantageous to this organism in rapidly adapting to favourable growth conditions.
BacchiC. J.,
MccannP. P.1987; Parasitic protozoa and polyamines. In Inhibition of Polyamine Metabolism pp. 317–344MccannP. P.,
PeggA. E.,
SjoerdsmaA.
Edited by San Diego: Academic Press;
FairlambA. H.,
CeramiA.1985; Identification of a novel thiol-containing co-factor essential for glutathione reductase in trypanosomatids. Molecular and Biochemical Parasitology 14:187–198
FairlambA. H.,
HendersonG. B.1987; Metabolism of trypanothione and glutathionylspermidine in trypanosomatids. In Host-Parasite Molecular Recognition and Interaction in Protozoal Infections pp. 29–40ChangK.-P.,
SnaryD.
Edited by Heidelberg: NATO ASI Series;
FairlambA. H.,
HendersonG. B.,
CeramiA.1986; The biosynthesis of trypanothione and N1- glutathionylspermidine in Crithidia fasciculata. Molecular and Biochemical Parasitology 21:247–257
FairlambA. H.,
HendersonG. B.,
BacchiC. J.,
CeramiA.1987; In vivo effects of difluoromethyl- omithine on trypanothione and polyamine levels in bloodstream forms of Trypanosoma brucei. Molecular and Biochemical Parasitology 24:185–191
HendersonG. B.,
UlrichP.,
FairlambA. H.,
CeramiA.1986; Synthesis of the trypanosomatid metabolites trypanothione, and N′-mono- and N8- mono-glutathionylspermidine. Journal of the Chemical Society Chemical-Communications593–594
Krauth-SiegelR. L.,
EndersB.,
HendersonG. B.,
FairlambA. H.,
SchirmerR. H.1987; Trypanothionereductase from Trypanosoma cruzi.Purification and characterization of the crystalline enzyme. European Journal of Biochemistry 164:123–128
TaborC. E.,
TaborH.1970; The complete conversion of spermidine to a peptide derivative in Escherichia coli. Biochemical and Biophysical Research Communications 41:232–238
TaborH.,
TaborC. E.1975; Isolation, characterization, and turnover of glutathionylspermidine from Escherichia coli. Journal of Biological Chemistry 250:2648–2654
ShamesS. L.,
FairlambA. H.,
CeramiA.,
WalshC. T.1986; Purification and characterization of trypanothionereductase from Crithidia fasciculata, a newly discovered member of the family of disulfide- containing flavoprotein reductases. Biochemistry 25:3519–3526
ZieglerD. M.1985; Role of reversible oxidation- reduction of enzyme thiols-disulfides in metabolic regulation. Annual Review of Biochemistry 54:305–329