scholarly journals Two-step phosphorylation of Ana2 by Plk4 is required for the sequential loading of Ana2 and Sas6 to initiate procentriole formation

Open Biology ◽  
2017 ◽  
Vol 7 (12) ◽  
pp. 170247 ◽  
Author(s):  
Nikola S. Dzhindzhev ◽  
George Tzolovsky ◽  
Zoltan Lipinszki ◽  
Mohammed Abdelaziz ◽  
Janus Debski ◽  
...  
Keyword(s):  
Active Form ◽  
Single Site ◽  
Outer Ring ◽  
Phosphorylation Sites ◽  
Serine Residue ◽  
Centriole Duplication ◽  
Catalytically Active ◽  
Zone Ii ◽  
Conserved Region ◽  
A Site

The conserved process of centriole duplication requires Plk4 kinase to recruit and promote interactions between Sas6 and Sas5/Ana2/STIL (respective nomenclature of worms/flies/humans). Plk4-mediated phosphorylation of Ana2/STIL in its conserved STAN motif has been shown to promote its interaction with Sas6. However, STAN motif phosphorylation is not required for recruitment of Ana2 to the centriole. Here we show that in Drosophila , Ana2 loads onto the site of procentriole formation ahead of Sas6 in a process that also requires Plk4. However, whereas Plk4 is first recruited to multiple sites around the ring of zone II at the periphery of the centriole, Ana2 is recruited to a single site in telophase before Plk4 becomes finally restricted to this same single site. When we over-ride the auto-destruction of Plk4, it remains localized to multiple sites in the outer ring of the centriole and, if catalytically active, recruits Ana2 to these sites. Thus, it is the active form of Plk4 that promotes Ana2's recruitment to the centriole. We now show that Plk4 phosphorylates Ana2 at a site other than the STAN motif, which lies in a conserved region we term the ANST (ANa2-STil) motif. Mutation of this site, S38, to a non-phosphorylatable residue prevents the procentriole loading of Ana2 and blocks centriole duplication. Thus the initiation of procentriole formation requires Plk4 to first phosphorylate a single serine residue in the ANST motif to promote Ana2's recruitment and, secondly, to phosphorylate four residues in the STAN motif enabling Ana2 to recruit Sas6. We discuss these findings in light of the multiple Plk4 phosphorylation sites on Ana2.

scholarly journals Pathways in the activation of human coagulation factor X

1980 ◽  
Vol 185 (3) ◽  
pp. 647-658 ◽  
Author(s):  
K Mertens ◽  
R M Bertina
Keyword(s):  
Heavy Chain ◽  
Active Form ◽  
Factor Xa ◽  
Coagulation Factor ◽  
Terminal Region ◽  
Factor Vii ◽  
Factor X ◽  
Catalytically Active ◽  
A Site ◽  
Polypeptide Chains

Purified human Factor X (apparent mol.wt. 72000), which consists of two polypeptide chains (mol.wt. 55000 and 19000), was activated by both Russell's-viper venom and the purified physiological activators (Factor VII/tissue factor and Factor IXa/Factor VIII). They all convert Factor X to catalytically active Factor Xa (mol.wt. 54000) by cleaving the heavy chain at a site on the N-terminal region. In the presence of Ca2+ and phospholipid, the Factor Xa formed catalyses (a) the cleavage of a small peptide (mol.wt. 4000) from the C-terminal region of the heavy chain of Factor Xa, resulting in a second active form (mol.wt. 50000), and (b) the cleavage of a peptide containing the active-site serine residue (mol.wt. 13000) from the C-terminal region of the heavy chain of Factor X, resulting in an inactivatable component (mol.wt. 59000). A nomenclature for the various products is proposed.

scholarly journals Thermal inactivation and chaperonin-mediated renaturation of mitochondrial aspartate aminotransferase

1998 ◽  
Vol 334 (1) ◽  
pp. 219-224 ◽  
Author(s):  
James M. LAWTON ◽  
Shawn DOONAN
Keyword(s):  
Escherichia Coli ◽  
Aspartate Aminotransferase ◽  
Thermal Inactivation ◽  
Active Form ◽  
Active Conformation ◽  
E Coli ◽  
Catalytically Active ◽  
Mitochondrial Aspartate Aminotransferase ◽  
Chaperonin 10 ◽  
Chaperonin 60

Mitochondrial aspartate aminotransferase is inactivated irreversibly on heating. The inactivated protein aggregates, but aggregation is prevented by the presence of the chaperonin 60 from Escherichia coli (GroEL). The chaperonin increases the rate of thermal inactivation in the temperature range 55–65 °C but not at lower temperatures. It has previously been shown [Twomey and Doonan (1997) Biochim. Biophys. Acta 1342, 37–44] that the enzyme switches to a modified, but catalytically active, conformation at approx. 55–60 °C and the present results show that this conformation is recognized by and binds to GroEL. The thermally inactivated protein can be released from GroEL in an active form by the addition of chaperonin 10 from E. coli (GroES)/ATP, showing that inactivation is not the result of irreversible chemical changes. These results suggest that the irreversibility of thermal inactivation is due to the formation of an altered conformation with a high kinetic barrier to refolding rather than to any covalent changes. In the absence of chaperonin the unfolded molecules aggregate but this is a consequence, rather than the cause, of irreversible inactivation.

scholarly journals Chiral Chalcogen Bond Donors Based on the 4,4′-Bipyridine Scaffold

Molecules ◽  
2019 ◽  
Vol 24 (24) ◽  
pp. 4484 ◽  
Author(s):  
Robin Weiss ◽  
Emmanuel Aubert ◽  
Paola Peluso ◽  
Sergio Cossu ◽  
Patrick Pale ◽  
...  
Keyword(s):  
High Performance Liquid Chromatography ◽  
High Performance ◽  
Chiral Stationary Phase ◽  
Asymmetric Induction ◽  
Selenium Compound ◽  
Design And Synthesis ◽  
Catalytically Active ◽  
A Site ◽  
Site Selective ◽  
Proof Of Principle

Organocatalysis through chalcogen bonding (ChB) is in its infancy, as its proof-of-principle was only reported in 2016. Herein, we report the design and synthesis of new chiral ChB donors, as well as the catalytic activity evaluation of the 5,5′-dibromo-2,2′-dichloro-3-((perfluorophenyl)selanyl)-4,4′-bipyridine as organocatalyst. The latter is based on the use of two electron-withdrawing groups, a pentafluorophenyl ring and a tetrahalo-4,4′-bipyridine skeleton, as substituents at the selenium center. Atropisomery of the tetrahalo-4,4′-bipyridine motif provides a chiral environment to these new ChB donors. Their synthesis was achieved through either selective lithium exchange and trapping or a site-selective copper-mediated reaction. Pure enantiomers of the 3-selanyl-4,4′-bipyridine were obtained by high performance liquid chromatography enantioseparation on specific chiral stationary phase, and their absolute configuration was assigned by comparison of the measured and calculated electronic circular dichroism spectra. The capability of the selenium compound to participate in σ-hole-based interactions in solution was studied by 19F NMR. Even if no asymmetric induction has been observed so far, the new selenium motif proved to be catalytically active in the reduction of 2-phenylquinoline by Hantzsch ester.

On the roles of calcium and zinc ions in the formation of a catalytically active form of the metalloenzyme, l-alanyl-d-glutamate peptidase of the bacteriophage T5 (EndoT5)

2020 ◽  
Vol 164 ◽  
pp. 2711-2716 ◽  
Author(s):  
Victor P. Kutyshenko ◽  
Galina V. Mikoulinskaia ◽  
Dmitry A. Prokhorov ◽  
Nikolai V. Molochkov ◽  
Alexander Y. Yegorov ◽  
...  
Keyword(s):  
Active Form ◽  
Zinc Ions ◽  
Catalytically Active ◽  
Bacteriophage T5

scholarly journals The Anti-Tumor Agent Sodium Selenate Decreases Methylated PP2A, Increases GSK3βY216 Phosphorylation, Including Tau Disease Epitopes and Reduces Neuronal Excitability in SHSY-5Y Neurons

2019 ◽  
Vol 20 (4) ◽  
pp. 844 ◽  
Author(s):  
Wesal Habbab ◽  
Imad Aoudé ◽  
Freshteh Palangi ◽  
Sara Abdulla ◽  
Tariq Ahmed
Keyword(s):  
Neurological Disorders ◽  
Neuronal Excitability ◽  
Active Form ◽  
Close Correlation ◽  
Maximum Effect ◽  
Sodium Selenate ◽  
Dependent Manner ◽  
Maximum Increase ◽  
Electrophysiological Studies ◽  
Catalytically Active

Selenium application as sodium selenate was repeatedly shown to have anti-carcinogenic properties by increasing levels of the serine/ threonine protein phosphatase 2A (PP2A) in cancer cells. PP2A has a prominent role in cell development, homeostasis, and in neurons regulates excitability. PP2A, GSK3β and Tau reside together in a complex, which facilitates their interaction and (dys)-function as has been reported for several neurological disorders. In this study we recorded maximum increase in total PP2A at 3 µM sodium selenate in a neuron cell line. In conjunction with these data, whole-cell electrophysiological studies revealed that this concentration had maximum effect on membrane potentials, conductance and currents. Somewhat surprisingly, the catalytically active form, methylated PP2A (mePP2A) was significantly decreased. In close correlation to these data, the phosphorylation state of two substrate proteins, sensitive to PP2A activity, GSK3β and Tau were found to be increased. In summary, our data reveal that sodium selenate enhances PP2A levels, but reduces catalytic activity of PP2A in a dose dependent manner, which fails to reduce Tau and GSK3β phosphorylation under physiological conditions, indicating an alternative route in the rescue of cell pathology in neurological disorders.

scholarly journals The missing link: allostery and catalysis in the anti-viral protein SAMHD1

2019 ◽  
Vol 47 (4) ◽  
pp. 1013-1027 ◽  
Author(s):  
Elizabeth R. Morris ◽  
Ian A. Taylor
Keyword(s):  
Allosteric Regulation ◽  
Active Form ◽  
Inflammatory Disorder ◽  
Dna Viruses ◽  
Allosteric Coupling ◽  
Allosteric Sites ◽  
Catalytically Active ◽  
Important Regulator ◽  
Hydrolysis Of ◽  
Vertebrate Protein

Abstract Vertebrate protein SAMHD1 (sterile-α-motif and HD domain containing protein 1) regulates the cellular dNTP (2′-deoxynucleoside-5′-triphosphate) pool by catalysing the hydrolysis of dNTP into 2′-deoxynucleoside and triphosphate products. As an important regulator of cell proliferation and a key player in dNTP homeostasis, mutations to SAMHD1 are implicated in hypermutated cancers, and germline mutations are associated with Chronic Lymphocytic Leukaemia and the inflammatory disorder Aicardi–Goutières Syndrome. By limiting the supply of dNTPs for viral DNA synthesis, SAMHD1 also restricts the replication of several retroviruses, such as HIV-1, and some DNA viruses in dendritic and myeloid lineage cells and resting T-cells. SAMHD1 activity is regulated throughout the cell cycle, both at the level of protein expression and post-translationally, through phosphorylation. In addition, allosteric regulation further fine-tunes the catalytic activity of SAMHD1, with a nucleotide-activated homotetramer as the catalytically active form of the protein. In cells, GTP and dATP are the likely physiological activators of two adjacent allosteric sites, AL1 (GTP) and AL2 (dATP), that bridge monomer–monomer interfaces to stabilise the protein homotetramer. This review summarises the extensive X-ray crystallographic, biophysical and molecular dynamics experiments that have elucidated important features of allosteric regulation in SAMHD1. We present a comprehensive mechanism detailing the structural and protein dynamics components of the allosteric coupling between nucleotide-induced tetramerization and the catalysis of dNTP hydrolysis by SAMHD1.

Variability in nitrogen content of submerged aquatic vegetation: utility as an indicator of N dynamics within and among lakes

2012 ◽  
Vol 65 (7) ◽  
pp. 1151-1157 ◽  
Author(s):  
Catherine Blanchet ◽  
Gabriel Maltais-Landry ◽  
Roxane Maranger
Keyword(s):  
Nitrogen Content ◽  
Plant Species ◽  
Aquatic Ecosystems ◽  
Submerged Aquatic Vegetation ◽  
Aquatic Vegetation ◽  
Spatial Scales ◽  
Single Site ◽  
N Availability ◽  
N Dynamics ◽  
A Site

Submerged aquatic vegetation (SAV) may serve as an integrative proxy of spatial and temporal nitrogen (N) availability in aquatic ecosystems as plants are physiologically capable of storing variable amounts of N. However, it is important to understand whether plant species behave similarly or differently within and among systems. We sampled different SAV species along a nutrient gradient at multiple sites within several lakes to determine variability in C:N ratios and % N content among species, among plants of the same species at a single site, among sites and among lakes. Species respond differently suggesting that not all plant types can be used universally as nutrient proxies. The greatest variability in % N and C:N ratios for Valliseneria americana was observed among lakes whereas for Elodea canadensis it was among sites within a lake and among plants within a site. This suggests that V. americana could be a particularly useful indicator of N availability at larger spatial scales (regional and within a large fluvial lake) but that E. canadensis was not a particularly useful proxy.

scholarly journals Activation and selectivity of OTUB-1 and OTUB-2 deubiquitinylases

2020 ◽  
Vol 295 (20) ◽  
pp. 6972-6982
Author(s):  
Dakshinamurthy Sivakumar ◽  
Vikash Kumar ◽  
Michael Naumann ◽  
Matthias Stein
Keyword(s):  
Active Site ◽  
Electrostatic Interactions ◽  
Active Form ◽  
Cysteine Proteases ◽  
Binding Pocket ◽  
Catalytic Triad ◽  
Catalytic Site ◽  
Dynamics Simulation ◽  
Active Site Residues ◽  
Catalytically Active

The ovarian tumor domain (OTU) deubiquitinylating cysteine proteases OTUB1 and OTUB2 (OTU ubiquitin aldehyde binding 1 and 2) are representative members of the OTU subfamily of deubiquitinylases. Deubiquitinylation critically regulates a multitude of important cellular processes, such as apoptosis, cell signaling, and growth. Moreover, elevated OTUB expression has been observed in various cancers, including glioma, endometrial cancer, ovarian cancer, and breast cancer. Here, using molecular dynamics simulation approaches, we found that both OTUB1 and OTUB2 display a catalytic triad characteristic of proteases but differ in their configuration and protonation states. The OTUB1 protein had a prearranged catalytic site, with strong electrostatic interactions between the active-site residues His265 and Asp267. In OTUB2, however, the arrangement of the catalytic triad was different. In the absence of ubiquitin, the neutral states of the catalytic-site residues in OTUB2 were more stable, resulting in larger distances between these residues. Only upon ubiquitin binding did the catalytic triad in OTUB2 rearrange and bring the active site into a catalytically feasible state. An analysis of water access channels revealed only a few diffusion trajectories for the catalytically active form of OTUB1, whereas in OTUB2 the catalytic site was solvent-accessible, and a larger number of water molecules reached and left the binding pocket. Interestingly, in OTUB2, the catalytic residues His224 and Asn226 formed a stable hydrogen bond. We propose that the observed differences in activation kinetics, protonation states, water channels, and active-site accessibility between OTUB1 and OTUB2 may be relevant for the selective design of OTU inhibitors.

Spatial epidemics with large finite range

1996 ◽  
Vol 33 (4) ◽  
pp. 933-939 ◽  
Author(s):  
Mathew D. Penrose
Keyword(s):  
Single Site ◽  
Finite Range ◽  
Total Rate ◽  
Critical Rate ◽  
A Site ◽  
Spatial Epidemics

In the epidemic with removal with range r, each site z, once infected, remains so for a period of time Tz, the variables Tz being i.i.d. with mean μ. While infected, a site infects its healthy r-neighbours independently at total rate α. After infection, sites become immune. We show that the critical rate of infection αc (r), above which an epidemic starting from a single site may continue forever, converges to μ–1 as r →∞.

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