Structural Insight into Activation Mechanism of Toxoplasma gondii Nucleoside Triphosphate Diphosphohydrolases by Disulfide Reduction

Krug, Ulrike; Zebisch, M.; Krauss, Michel; Sträter, Norbert

doi:10.1074/jbc.m111.294348

Cited by 22 publications

(37 citation statements)

References 54 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In the NTPDases of the protozoan parasite Toxoplasma gondii, a restriction of a 12°domain closure motion is used to retain the enzyme in an inactive form. Reduction of a regulatory disulfide bridge initiates a domain closure and leads to activity [199].…”

Section: Active Site and Catalytic Mechanismmentioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

Self Cite

888

922

View full text Add to dashboard Cite

Ecto-nucleotidases play a pivotal role in purinergic signal transmission. They hydrolyze extracellular nucleotides and thus can control their availability at purinergic P2 receptors. They generate extracellular nucleosides for cellular reuptake and salvage via nucleoside transporters of the plasma membrane. The extracellular adenosine formed acts as an agonist of purinergic P1 receptors. They also can produce and hydrolyze extracellular inorganic pyrophosphate that is of major relevance in the control of bone mineralization. This review discusses and compares four major groups of ectonucleotidases: the ecto-nucleoside triphosphate diphosphohydrolases, ecto-5′-nucleotidase, ecto-nucleotide pyrophosphatase/phosphodiesterases, and alkaline phosphatases. Only recently and based on crystal structures, detailed information regarding the spatial structures and catalytic mechanisms has become available for members of these four ecto-nucleotidase families. This permits detailed predictions of their catalytic mechanisms and a comparison between the individual enzyme groups. The review focuses on the principal biochemical, cell biological, catalytic, and structural properties of the enzymes and provides brief reference to tissue distribution, and physiological and pathophysiological functions.

show abstract

Section: Active Site and Catalytic Mechanismmentioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

Self Cite

888

922

View full text Add to dashboard Cite

show abstract

“…In the structure of Tg NTPDase3 wt the active site cleft is open and the catalytic residues are not properly positioned for the hydrolysis of nucleotides. Variants of Tg NTPDase1 (PDB: 4A5B) and ‐3 (PDB: 4A5A) lacking the C258/C268 disulfide bridge (in the following text designated by the symbol “ΔCC”) indicated that disulfide reduction results in a conformational change of the activation loop in which the disulfide is located, causing (i) a 12 Å contraction of the tetramer along the molecular r‐dyad, (ii) a ∼13° rotation of the two dimers within the tetramer around the r‐axis and (iii) an 11° closure motion of the two domains in each monomer. Thereby the active site residues are arranged to a catalytically competent formation as observed in other NTPDases …”

Section: Introductionmentioning

confidence: 92%

“…The two NTPDases differ only by 16 out of 628 amino acids, but show large differences in their ability to hydrolyze nucleotides. NTPDase3 shows a strong preference for ATP over ADP while NTPDase1 hydrolyzes ATP and ADP about equally well . Interestingly, the enzymes need to be activated to be able to hydrolyze nucleoside triphosphates and –diphosphates .…”

Section: Introductionmentioning

confidence: 99%

“…Interestingly, the enzymes need to be activated to be able to hydrolyze nucleoside triphosphates and –diphosphates . It was shown that the activation is related to the reduction of the disulfide bond C258/C268 …”

Section: Introductionmentioning

confidence: 99%

“…Crystal structures of T. gondii NTPDase3 wild‐type ( Tg NTPDase3 wt, PDBs: 4A57 and 4A59) revealed a homotetrameric structure of D 2 symmetry . In the structure of Tg NTPDase3 wt the active site cleft is open and the catalytic residues are not properly positioned for the hydrolysis of nucleotides.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

The crystal structure of Toxoplasma gondii nucleoside triphosphate diphosphohydrolase 1 represents a conformational intermediate in the reductive activation mechanism of the tetrameric enzyme

2013

Self Cite

View full text Add to dashboard Cite

Toxoplasma gondii nucleoside triphosphate diphosphohydrolase (NTPDase) 1 was crystallized in an intermediate tetrameric conformation. The crystal structure is similar to that of T. gondii NTPDase3 and represents an inactive conformation as the activating disulfide bridge is not reduced and the active site cleft between the two domains of each monomer is open. However, the arrangement of the monomers within the tetramer differs from that of the inactive form of NTPDase3 and may represent an intermediate conformation on the path of the closure motion of the tetramer induced upon activation.

show abstract

The ATP/ADP Substrate Specificity Switch between Toxoplasma gondii NTPDase1 and NTPDase3 is Caused by an Altered Mode of Binding of the Substrate Base

Krug¹,

Totzauer²,

Zebisch³

et al. 2013

ChemBioChem

View full text Add to dashboard Cite

Two nucleoside triphosphate diphosphohydrolase isoforms (NTPDase1 and NTPDase3) are responsible for the hydrolysis of nucleotides by the intracellular protozoan Toxoplasma gondii. They constitute about 3 % of the total parasite protein. Despite sharing 97 % sequence identity they exhibit opposite ATP versus ADP substrate discrimination ratios. Here we show by mutagenesis that the residues G492/G493 in NTPDase3 and R492/E493 in NTPDase1 are predominantly responsible for the differences in substrate specificity. Crystal structures of NTPDase1 in complexation with analogues of ATP and ADP reveal that the inverted substrate specificity of NTPDase1 relative to NTPDase3 is achieved by switching from the canonical substrate binding mode to a very different alternative one. Instead of being stacked on top of a helix of the C-terminal domain the nucleotide base is positioned in the interdomain space between the side chains of R108 and R492, recruited from both domains. Furthermore, we show that the NTPDase1 substrate specificity is mainly dependent on the presence of the side chain of E493, which causes repositioning of the ribose component of the nucleotide. All in all, binding by the flexible side chains in the alternative binding mode in NTPDase1 allows for equally good positioning of ATP and ADP with increased activity toward ADP relative to what is seen in the case of NTPDase3.

show abstract

Structural Insight into Activation Mechanism of Toxoplasma gondii Nucleoside Triphosphate Diphosphohydrolases by Disulfide Reduction

Cited by 22 publications

References 54 publications

Cellular function and molecular structure of ecto-nucleotidases

Cellular function and molecular structure of ecto-nucleotidases

The crystal structure of Toxoplasma gondii nucleoside triphosphate diphosphohydrolase 1 represents a conformational intermediate in the reductive activation mechanism of the tetrameric enzyme

The ATP/ADP Substrate Specificity Switch between Toxoplasma gondii NTPDase1 and NTPDase3 is Caused by an Altered Mode of Binding of the Substrate Base

Contact Info

Product

Resources

About