Literature summary for 2.4.2.8 extracted from

Valsecchi, W.M.; Cousido-Siah, A.; Defelipe, L.A.; Mitschler, A.; Podjarny, A.; Santos, J.; Delfino, J.M.
The role of the C-terminal region on the oligomeric state and enzymatic activity of Trypanosoma cruzi hypoxanthine phosphoribosyl transferase (2016), Biochim. Biophys. Acta, 1864, 655-666 .

Application

Application	Comment	Organism
drug development	the enzyme is a possible target for anti-parasite drug development	Trypanosoma cruzi

Cloned(Commentary)

Cloned (Comment)	Organism
gene HGPRTase, recombinant expression of His6-tagged enzyme and of nontagged enzyme in Escherichia coli strain BL21(DE3)	Trypanosoma cruzi

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant His6-tagged enzyme, mixing of 5 mg/ml protein in 20 mM Tris-HCl, 100 mM NaCl, pH 8.0, with reservoir solution containing 0.1 M MES, pH 6.5, and 12% w/v PEG 20000 at 20°C, X-ray diffraction structure determination and analysis at 2.65 A resolution	Trypanosoma cruzi

Protein Variants

Protein Variants	Comment	Organism
additional information	truncation of the C-terminal stretch of enzyme TcHPRTH6 enhances its catalytic efficiency	Trypanosoma cruzi

General Stability

General Stability	Organism
analysis of the stability of the variants in urea-induced unfolding experiments. Protein aggregation is taking place after dialysis or denaturant dilution, an indication of the existence of an irreversible phenomenon, precludes the derivation of thermodynamic state functions from the unfolding data. Nevertheless, the enzyme behaves as cooperative folding units when submitted to chemical denaturation, irreversible aggregation at 60°C within 10 min	Trypanosoma cruzi

Organism

analysis of the stability of the variants in urea-induced unfolding experiments. Protein aggregation is taking place after dialysis or denaturant dilution, an indication of the existence of an irreversible phenomenon, precludes the derivation of thermodynamic state functions from the unfolding data. Nevertheless, the enzyme behaves as cooperative folding units when submitted to chemical denaturation, irreversible aggregation at 60°C within 10 min

Trypanosoma cruzi

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	required	Trypanosoma cruzi

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
hypoxanthine + 5-phospho-alpha-D-ribose 1-diphosphate	Trypanosoma cruzi	-	IMP + diphosphate	-	?

Organism

Organism	UniProt	Comment	Textmining
Trypanosoma cruzi	Q27796	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His6-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and dialysis, recombinant nontagged enzyme from Escherichia coli by anion exchange chromatography, ammonium sulfate fractionation, dialysis, and gel filtration, both to over 95% purity	Trypanosoma cruzi

Renatured (Commentary)

Renatured (Comment)	Organism
although the enzyme protein irreversibly aggregates during unfolding, oligomerization is reversible and can be modulated by low concentrations of urea	Trypanosoma cruzi

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
hypoxanthine + 5-phospho-alpha-D-ribose 1-diphosphate	-	Trypanosoma cruzi	IMP + diphosphate	-	?

Subunits

Subunits	Comment	Organism
dimer	when the C-terminal region, which is predicted as a disordered stretch, is excised by proteolysis, TcHPRT adopts a dimeric state, suggesting that the C-terminal region acts as a main guide for the quaternary arrangement. The C-terminal region exhibits a modulatory role on the enzyme, as attested by the enhanced activity observed for the dimeric form	Trypanosoma cruzi
tetramer	the full-length enzyme form in solution adopts a stable and enzymatically active tetrameric form, exhibiting large intersubunit surfaces, two putative dimeric interfaces	Trypanosoma cruzi

Synonyms

Synonyms	Comment	Organism
HGPRTase	-	Trypanosoma cruzi
hypoxanthine phosphoribosyl transferase	-	Trypanosoma cruzi
TcHPRT	-	Trypanosoma cruzi

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Trypanosoma cruzi

Temperature Stability [°C]

Temperature Stability Minimum [°C]	Temperature Stability Maximum [°C]	Comment	Organism
60	-	analysis of the stability of the variants in temperature unfolding experiments. Protein aggregation is taking place after dialysis or denaturant dilution, an indication of the existence of an irreversible phenomenon, precludes the derivation of thermodynamic state functions from the unfolding data. But the enzyme behave as cooperative folding units when submitted to thermal denaturation. In particular, a qualitative comparison (a shift in the observed apparent Tm values) of thermal denaturation curves suggests that the His-tag present in TcHPRTH6 might enhance its thermostability by increasing the height of the activation barrier to the formation of aggregation-prone species. Nevertheless, when the enzyme is incubated for 10 min at 60°C, it irreversibly aggregates with a concomitant loss of enzymatic activity	Trypanosoma cruzi

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.4	-	assay at	Trypanosoma cruzi

General Information

General Information	Comment	Organism
metabolism	the metabolism of the protozoan parasite Trypanosoma cruzi depends on the salvage of exogenous purines for nucleotide synthesis. In this context, TcHPRT plays a key role in the survival of trypanosomes in their hosts	Trypanosoma cruzi
physiological function	the hypoxanthine phosphoribosyl transferase (TcHPRT) is a critical enzyme in Trypanosoma cruzi for the survival of the parasite. TcHPRT catalyzes the transfer of a mono phosphorylated ribose from phosphoribosyl diphosphate (PRPP) to the purine ring	Trypanosoma cruzi