Ribonucleotide reductase is regulated via the R2 subunit during the life cycle of <i>Trypanosoma brucei</i>

Breidbach, Tanja; Krauth‐Siegel, R. Luise; Steverding, Dietmar

doi:10.1016/s0014-5793(00)01533-7

Cited by 12 publications

(15 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Taking into account the need for equimolar amounts of both subunits to form an active holoenzyme, nrdHIE transcripts turn out to be the limiting factor for ribonucleotide reduction in C. ammoniagenes. Nevertheless, when the expression of a gene is studied at the mRNA level, it must be borne in mind that post-transcriptional or other generegulatory mechanisms may be present; this has been shown to be of great importance in mammalian and malaria parasite RNRs (Björklund et al, 1990;Breidbach et al, 2000).…”

Section: Discussionmentioning

confidence: 99%

Corynebacterium ammoniagenes class Ib ribonucleotide reductase: transcriptional regulation of an atypical genomic organization in the nrd cluster

2003

View full text Add to dashboard Cite

Section: Discussionmentioning

confidence: 99%

Corynebacterium ammoniagenes class Ib ribonucleotide reductase: transcriptional regulation of an atypical genomic organization in the nrd cluster

2003

View full text Add to dashboard Cite

“…RNAi cell lines were maintained in the presence of the selectable agents hygromycin (50 g/ml), G-418 (15 g/ml), and phleomycin (3 g/ml) until 48 h prior to the start of an experiment. Extracts from long slender and short stumpy bloodstream trypanosomes were prepared and analyzed using antibody markers as described previously (21). Cell counts were determined using either a Neubauer heamocytometer (Sigma), or a CASY1 cell counter (Schä rfe System).…”

Section: Methodsmentioning

confidence: 99%

“…Polyclonal rabbit antibodies raised against T. brucei aldolase were used as described previously (27). SDS-PAGE and immunoblotting were carried out according to standard methods (21). Immunofluorescence of formaldehyde-and methanol-fixed detergent-extracted cytoskeletons using BB2 and ROD-1 (28) monoclonal antibodies was performed as described previously (29,30).…”

Section: Methodsmentioning

confidence: 99%

“…A lower amount of ADKG was found in the cell cycle-arrested bloodstream stumpy forms (19% of the level seen in long slender pleomorphic AnTat 1.1 bloodstream forms; 28% of the level seen in procyclic trypanosomes) that are preadapted for rapid differentiation into procyclic forms in the tsetse fly midgut, indicating some degree of stage-specific regulation. The homogeneity of such parasite populations was confirmed using antibodies against known stage-regulated proteins, as described previously (21).…”

Section: Seven Adk-like Genes Are Expressed In T Brucei-usingmentioning

confidence: 99%

See 1 more Smart Citation

Intracellular Positioning of Isoforms Explains an Unusually Large Adenylate Kinase Gene Family in the Parasite Trypanosoma brucei

Ginger

Ngazoa

Pereira

et al. 2005

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Adenylate kinases occur classically as cytoplasmic and mitochondrial enzymes, but the expression of seven adenylate kinases in the flagellated protozoan parasite Trypanosoma brucei (order, Kinetoplastida; family, Trypanosomatidae) easily exceeds the number of isoforms previously observed within a single cell and raises questions as to their location and function. We show that a requirement to target adenylate kinase into glycosomes, which are unique kinetoplastid-specific microbodies of the peroxisome class in which many reactions of carbohydrate metabolism are compartmentalized, and two different flagellar structures as well as cytoplasm and mitochondrion explains the expansion of this gene family in trypanosomes. The three isoforms that are selectively built into either the flagellar axoneme or the extra-axonemal paraflagellar rod, which is essential for motility, all contain long N-terminal extensions. Biochemical analysis of the only short form trypanosome adenylate kinase revealed that this enzyme catalyzes phosphotransfer of ␥-phosphate from ATP to AMP, CMP, and UMP acceptors; its high activity and specificity toward CMP is likely to reflect an adaptation to very low intracellular cytidine nucleotide pools. Analysis of some of the phosphotransfer network using RNA interference suggests considerable complexity within the homeostasis of cellular energetics. The anchoring of specific adenylate kinases within two distinct flagellar structures provides a paradigm for metabolic organization and efficiency in other flagellates.

show abstract

“…T. brucei ribonucleotide reductase is regulated via the R2 subunit. Whereas the R1 protein is present throughout the life cycle of the parasite, the R2 protein is not found in cell cycle-arrested short stumpy trypanosomes (6).…”

mentioning

confidence: 96%

Trypanothione-dependent Synthesis of Deoxyribonucleotides by Trypanosoma brucei Ribonucleotide Reductase

Dormeyer¹,

Reckenfelderbäumer

Lüdemann

et al. 2001

Journal of Biological Chemistry

113

View full text Add to dashboard Cite

Trypanosoma brucei, the causative agent of African sleeping sickness, synthesizes deoxyribonucleotides via a classical eukaryotic class I ribonucleotide reductase. The unique thiol metabolism of trypanosomatids in which the nearly ubiquitous glutathione reductase is replaced by a trypanothione reductase prompted us to study the nature of thiols providing reducing equivalents for the parasite synthesis of DNA precursors. Here we show that the dithiol trypanothione (bis(glutathionyl)spermidine), in contrast to glutathione, is a direct reductant of T. brucei ribonucleotide reductase with a K m value of 2 mM. This is the first example of a natural low molecular mass thiol directly delivering reducing equivalents for ribonucleotide reduction. At submillimolar concentrations, the reaction is strongly accelerated by tryparedoxin, a 16-kDa parasite protein with a WCPPC active site motif. The K m value of T. brucei ribonucleotide reductase for T. brucei tryparedoxin is about 4 M. The disulfide form of trypanothione is a powerful inhibitor of the tryparedoxin-mediated reaction that may represent a physiological regulation of deoxyribonucleotide synthesis by the redox state of the cell. The trypanothione/tryparedoxin system is a new system providing electrons for a class I ribonucleotide reductase, in addition to the well known thioredoxin and glutaredoxin systems described in other organisms.Ribonucleotide reductases (E.C. 1.17.4.1) catalyze the ratelimiting step in the de novo synthesis of DNA precursors and thus are key enzymes for the replication of an organism (1). African trypanosomes possess a typical eukaryotic class I ribonucleotide reductase (2-4). The genes encoding the Trypanosoma brucei R1 and R2 1 proteins have been cloned and overexpressed in Escherichia coli. Class I enzymes are tetrameric proteins composed of two R1 and R2 molecules each. The large R1 protein harbors the active site, as well as regulatory sites, and the small R2 protein contains a -oxo-bridged diiron cluster, which represents the Fe(III)-O 2Ϫ -Fe(III) cofactor in all eukaryotic ribonucleotide reductases, and a tyrosyl radical essential for catalysis (1, 5). T. brucei ribonucleotide reductase is regulated via the R2 subunit. Whereas the R1 protein is present throughout the life cycle of the parasite, the R2 protein is not found in cell cycle-arrested short stumpy trypanosomes (6

show abstract

Ribonucleotide reductase is regulated via the R2 subunit during the life cycle of Trypanosoma brucei

Cited by 12 publications

References 34 publications

Corynebacterium ammoniagenes class Ib ribonucleotide reductase: transcriptional regulation of an atypical genomic organization in the nrd cluster

Corynebacterium ammoniagenes class Ib ribonucleotide reductase: transcriptional regulation of an atypical genomic organization in the nrd cluster

Intracellular Positioning of Isoforms Explains an Unusually Large Adenylate Kinase Gene Family in the Parasite Trypanosoma brucei

Trypanothione-dependent Synthesis of Deoxyribonucleotides by Trypanosoma brucei Ribonucleotide Reductase

Contact Info

Product

Resources

About