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Biopolymers Aug 2016Microtubules are amazing filaments made of GTPase enzymes that store energy used for their own self-destruction to cause a stochastically driven dynamics called dynamic... (Review)
Review
Microtubules are amazing filaments made of GTPase enzymes that store energy used for their own self-destruction to cause a stochastically driven dynamics called dynamic instability. Dynamic instability can be reproduced in vitro with purified tubulin, but the dynamics do not mimic that observed in cells. This is because stabilizers and destabilizers act to alter microtubule dynamics. One interesting and understudied class of destabilizers consists of the microtubule-severing enzymes from the ATPases Associated with various cellular Activities (AAA+) family of ATP-enzymes. Here we review current knowledge about GTP-driven microtubule dynamics and how that couples to ATP-driven destabilization by severing enzymes. We present a list of challenges regarding the mechanism of severing, which require development of experimental and modeling approaches to shed light as to how severing enzymes can act to regulate microtubule dynamics in cells. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 547-556, 2016.
Topics: Adenosine Triphosphatases; Animals; Humans; Microtubules; Pyrophosphatases; Tubulin; Tubulin Modulators
PubMed: 27037673
DOI: 10.1002/bip.22842 -
The Plant Cell May 2018
Topics: Arabidopsis; Diphosphates; Inorganic Pyrophosphatase; Pyrophosphatases; Vacuoles
PubMed: 29716989
DOI: 10.1105/tpc.18.00342 -
Clinical Pharmacology and Therapeutics May 2019
Topics: Alleles; Biotransformation; Cytochrome P-450 Enzyme System; Genome-Wide Association Study; Haplotypes; Humans; Medication Therapy Management; Pharmacogenetics; Pharmacogenomic Testing; Pharmacogenomic Variants; Pyrophosphatases; Terminology as Topic
PubMed: 30515762
DOI: 10.1002/cpt.1268 -
PLoS Pathogens Aug 2016
Review
Topics: Host-Pathogen Interactions; Plant Diseases; Plant Immunity; Plants; Pyrophosphatases; Nudix Hydrolases
PubMed: 27513453
DOI: 10.1371/journal.ppat.1005704 -
Scientific Reports Dec 2015ADP-ribosylation refers to the transfer of the ADP-ribose group from NAD(+) to target proteins post-translationally, either attached singly as mono(ADP-ribose) (MAR) or...
ADP-ribosylation refers to the transfer of the ADP-ribose group from NAD(+) to target proteins post-translationally, either attached singly as mono(ADP-ribose) (MAR) or in polymeric chains as poly(ADP-ribose) (PAR). Though ADP-ribosylation is therapeutically important, investigation of this protein modification has been limited by a lack of proteomic tools for site identification. Recent work has demonstrated the potential of a tag-based pipeline in which MAR/PAR is hydrolyzed down to phosphoribose, leaving a 212 Dalton tag at the modification site. While the pipeline has been proven effective by multiple groups, a barrier to application has become evident: the enzyme used to transform MAR/PAR into phosphoribose must be purified from the rattlesnake Crotalus adamanteus venom, which is contaminated with proteases detrimental for proteomic applications. Here, we outline the steps necessary to purify snake venom phosphodiesterase I (SVP) and describe two alternatives to SVP-the bacterial Nudix hydrolase EcRppH and human HsNudT16. Importantly, expression and purification schemes for these Nudix enzymes have already been proven, with high-quality yields easily attainable. We demonstrate their utility in identifying ADP-ribosylation sites on Poly(ADP-ribose) Polymerase 1 (PARP1) with mass spectrometry and discuss a structure-based rationale for this Nudix subclass in degrading protein-conjugated ADP-ribose, including both MAR and PAR.
Topics: Adenosine Diphosphate Ribose; Animals; Bacterial Proteins; Crotalid Venoms; Humans; Phosphodiesterase I; Proteomics; Pyrophosphatases; Viperidae; Nudix Hydrolases
PubMed: 26669448
DOI: 10.1038/srep18271 -
Angewandte Chemie (International Ed. in... Feb 2023(-)-Antrocin (1), produced by the medicinal mushroom Antrodia cinnamomea, is a potent antiproliferative compound. The biosynthetic gene cluster of 1 was identified, and...
(-)-Antrocin (1), produced by the medicinal mushroom Antrodia cinnamomea, is a potent antiproliferative compound. The biosynthetic gene cluster of 1 was identified, and the pathway was characterized by heterologous expression. We characterized a haloacid dehalogenase-like terpene cyclase AncC that biosynthesizes the drimane-type sesquiterpene (+)-albicanol (2) from farnesyl pyrophosphate (FPP). Biochemical characterization of AncC, including kinetic studies and mutagenesis, demonstrated the functions of two domains: a terpene cyclase (TC) and a pyrophosphatase (PPase). The TC domain first cyclizes FPP to albicanyl pyrophosphate, and the PPase domain then removes the pyrophosphate to form 2. Intriguingly, AncA (94 % sequence identity to AncC), in the same gene cluster, converts FPP into (R)-trans-γ-monocyclofarnesol instead of 2. Notably, Y283/F375 in the TC domain of AncA serve as a gatekeeper in controlling the formation of a cyclofarnesoid rather than a drimane-type scaffold.
Topics: Terpenes; Diphosphates; Agaricales; Antibodies, Antineutrophil Cytoplasmic; Kinetics; Sesquiterpenes; Pyrophosphatases; Multigene Family
PubMed: 36583947
DOI: 10.1002/anie.202215566 -
Journal of Orthopaedic Surgery and... Jun 2023The deficiency of ectonucleotide pyrophosphatase/phosphodiesterase 1 (Enpp1) causes the phenotype similar to knee osteoarthritis (OA). However, the molecular mechanism...
OBJECTIVE AND BACKGROUND
The deficiency of ectonucleotide pyrophosphatase/phosphodiesterase 1 (Enpp1) causes the phenotype similar to knee osteoarthritis (OA). However, the molecular mechanism is poorly understood.
METHOD
The global deletion of Enpp1 (Enpp1) mice was created to analyze the role of Enpp1 in the progress of knee OA. The apoptosis, proliferation and chondrogenic differentiation ability of chondrocytes from wild-type (WT) and Enpp1 joints were compared. According to the results of high-throughput quantitative molecular measurements, the proteins of chondrocytes from WT and Enpp1 mice were used to explore the mechanism of Enpp1 deficiency-associated knee OA.
RESULT
In Enpp1 knee joints, we found significant chondrocyte apoptosis and proteomic results showed that abnormal expression of AMP-activated protein kinase (AMPK) signaling pathway may contribute to this phenotype. In primary chondrocyte cultures in vitro, Enpp1 deletion dramatically enhancing chondrocyte apoptosis. Meanwhile, we found Enpp1 deletion inhibits the phosphorylation of AMPK (P-AMPK). We also found that decreased level of P-AMPK and chondrocyte apoptosis, which are caused by Enpp1 deficiency, can be reversed by Acadesine (AICAR), the activator of AMPK.
CONCLUSION
Consequently, Enpp1 deficiency plays an essential role in knee OA by regulating AMPK signaling pathway.
Topics: Animals; Mice; AMP-Activated Protein Kinases; Apoptosis; Chondrocytes; Osteoarthritis, Knee; Phosphoric Diester Hydrolases; Proteomics; Pyrophosphatases; Signal Transduction
PubMed: 37370114
DOI: 10.1186/s13018-023-03923-1 -
The FEBS Journal Sep 2016Deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase) is essential for genome integrity. Interestingly, this enzyme from Drosophila virilis has an unusual form, as...
Deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase) is essential for genome integrity. Interestingly, this enzyme from Drosophila virilis has an unusual form, as three monomer repeats are merged with short linker sequences, yielding a fused trimer-like dUTPase fold. Unlike homotrimeric dUTPases that are encoded by a single repeat dut gene copy, the three repeats of the D. virilis dut gene are not identical due to several point mutations. We investigated the potential evolutionary pathway that led to the emergence of this extant fused trimeric dUTPase in D. virilis. The herein proposed scenario involves two sequential gene duplications followed by sequence divergence amongst the dut repeats. This pathway thus requires the existence of a transient two-repeat-containing fused dimeric dUTPase intermediate. We identified the corresponding ancestral dUTPase single repeat enzyme together with its tandem repeat evolutionary intermediate and characterized their enzymatic function and structural stability. We additionally engineered and characterized artificial single or tandem repeat constructs from the extant enzyme form to investigate the influence of the emergent residue alterations on the formation of a functional assembly. The observed severely impaired stability and catalytic activity of these latter constructs provide a plausible explanation for evolutionary persistence of the extant fused trimeric D. virilis dUTPase form. For the ancestral homotrimeric and the fused dimeric intermediate forms, we observed strong catalytic and structural competence, verifying viability of the proposed evolutionary pathway. We conclude that the progression along the herein described evolutionary trajectory is determined by the retained potential of the enzyme for its conserved three-fold structural symmetry.
Topics: Amino Acid Sequence; Amino Acid Substitution; Animals; Drosophila; Drosophila Proteins; Enzyme Stability; Evolution, Molecular; Gene Duplication; Genes, Insect; Models, Molecular; Phylogeny; Point Mutation; Protein Folding; Protein Structure, Quaternary; Pyrophosphatases; Sequence Homology, Amino Acid; Tandem Repeat Sequences
PubMed: 27380921
DOI: 10.1111/febs.13800 -
The Plant Cell May 2018Inorganic pyrophosphate (PPi) is a phosphate donor and energy source. Many metabolic reactions that generate PPi are suppressed by high levels of PPi. Here, we...
Inorganic pyrophosphate (PPi) is a phosphate donor and energy source. Many metabolic reactions that generate PPi are suppressed by high levels of PPi. Here, we investigated how proper levels of cytosolic PPi are maintained, focusing on soluble pyrophosphatases (AtPPa1 to AtPPa5; hereafter PPa1 to PPa5) and vacuolar H-pyrophosphatase (H-PPase, AtVHP1/FUGU5) in In planta, five PPa isozymes tagged with GFP were detected in the cytosol and nuclei. Immunochemical analyses revealed a high abundance of PPa1 and the absence of PPa3 in vegetative tissue. In addition, the heterologous expression of each PPa restored growth in a soluble PPase-defective yeast strain. Although the quadruple knockout mutant plant showed no obvious phenotypes, H-PPase and PPa1 double mutants () exhibited significant phenotypes, including dwarfism, high PPi concentrations, ectopic starch accumulation, decreased cellulose and callose levels, and structural cell wall defects. Altered cell arrangements and weakened cell walls in the root tip were particularly evident in and were more severe than in Our results indicate that H-PPase is essential for maintaining adequate PPi levels and that the cytosolic PPa isozymes, particularly PPa1, prevent increases in PPi concentrations to toxic levels. We discuss phenotypes in relation to metabolic reactions and PPi homeostasis.
Topics: Arabidopsis; Cytosol; Diphosphates; Inorganic Pyrophosphatase; Pyrophosphatases; Vacuoles
PubMed: 29691313
DOI: 10.1105/tpc.17.00911 -
Journal of Oral Biosciences Sep 2021This study aimed to demonstrate the immunolocalization and gene expression of tissue nonspecific alkaline phosphatase (TNALP) and ectonucleotide...
OBJECTIVE
This study aimed to demonstrate the immunolocalization and gene expression of tissue nonspecific alkaline phosphatase (TNALP) and ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) in osteoblasts, preosteoblasts, and osteocytes of murine bone to provide clues for a better understanding of the supply of phosphate ions (Pi) during bone mineralization.
METHODS
Six-week-old male C57BL/6J mice (n = 6) were fixed with a paraformaldehyde solution, and the right femora were extracted for immunodetection of TNALP and ENPP1, while the left tibiae were used for reverse transcription polymerase chain reaction to evaluate Tnalp and Enpp1 gene expression.
RESULTS
TNALP was intensely localized on the basolateral cell membranes of mature osteoblasts and preosteoblastic cells. There was little immunoreactivity of TNALP on the secretory surface of the osteoblasts and no TNALP reactivity in the osteocytes. In contrast, ENPP1 was observed throughout the cytoplasm of mature osteoblasts and osteocytes embedded in bone but was not observed in preosteoblasts. Together, despite the fact that the osteoid is a site of matrix vesicle-mediated mineralization, ENPP1, which inhibits mineralization by providing pyrophosphates, was localized in close proximity of the osteoid, whereas TNALP, which facilitates mineralization by providing Pi, was relatively distant from the osteoid.
CONCLUSION
It seems likely that the differential localization of TNALP and ENPP1 around the osteoid observed at the microscopic level may provide preferential micro-circumstance for a balanced concentration of Pi and pyrophosphate for bone mineralization.
Topics: Alkaline Phosphatase; Animals; Male; Mice; Mice, Inbred C57BL; Osteocytes; Phosphoric Diester Hydrolases; Pyrophosphatases
PubMed: 34391947
DOI: 10.1016/j.job.2021.08.001