scholarly journals Cooperative Binding of the Cationic Porphyrin Tris-T4 Enhances Catalytic Activity of 20S Proteasome Unveiling a Complex Distribution of Functional States

2020 ◽  
Vol 21 (19) ◽  
pp. 7190
Author(s):  
Anna Maria Santoro ◽  
Alessandro D’Urso ◽  
Alessandra Cunsolo ◽  
Danilo Milardi ◽  
Roberto Purrello ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Interaction Model ◽  
In Silico Analysis ◽  
Cooperative Binding ◽  
20S Proteasome ◽  
Conformational Effects ◽  
Dynamic View ◽  
Functional States ◽  
New Generation ◽  
Substrate Hydrolysis

The present study provides new evidence that cationic porphyrins may be considered as tunable platforms to interfere with the structural “key code” present on the 20S proteasome α-rings and, by consequence, with its catalytic activity. Here, we describe the functional and conformational effects on the 20S proteasome induced by the cooperative binding of the tri-cationic 5-(phenyl)-10,15,20-(tri N-methyl-4-pyridyl) porphyrin (Tris-T4). Our integrated kinetic, NMR, and in silico analysis allowed us to disclose a complex effect on the 20S catalytic activity depending on substrate/porphyrin concentration. The analysis of the kinetic data shows that Tris-T4 shifts the relative populations of the multiple interconverting 20S proteasome conformations leading to an increase in substrate hydrolysis by an allosteric pathway. Based on our Tris-T4/h20S interaction model, Tris-T4 is able to affect gating dynamics and substrate hydrolysis by binding to an array of negatively charged and hydrophobic residues present on the protein surface involved in the 20S molecular activation by the regulatory proteins (RPs). Accordingly, despite the fact that Tris-T4 also binds to the α3ΔN mutant, allosteric modulation is not observed since the molecular mechanism connecting gate dynamics with substrate hydrolysis is impaired. We envisage that the dynamic view of the 20S conformational equilibria, activated through cooperative Tris-T4 binding, may work as a simplified model for a better understanding of the intricate network of 20S conformational/functional states that may be mobilized by exogenous ligands, paving the way for the development of a new generation of proteasome allosteric modulators.

CoOx–carbon nanotubes hybrids integrated on carbon cloth as a new generation of 3D porous hydrogen evolution promoters

2017 ◽  
Vol 5 (21) ◽  
pp. 10510-10516 ◽  
Author(s):  
Jing Wang ◽  
Zhongzhe Wei ◽  
Haiyan Wang ◽  
Yiqing Chen ◽  
Yong Wang
Keyword(s):  
Carbon Nanotubes ◽  
Catalytic Activity ◽  
Low Temperature ◽  
Hydrogen Evolution ◽  
Carbon Cloth ◽  
Wide Ph Range ◽  
Ph Range ◽  
New Generation

CoOx–CNT–CC electrodes not only displayed outstanding performance over a wide pH range, but also showed superb catalytic activity at low temperature.

scholarly journals Sortir de la grande nuit

Anos 90 ◽  
2014 ◽  
Vol 21 (40) ◽  
Author(s):  
Adriano Moraes Migliavacca
Keyword(s):  
United States Of America ◽  
The United States ◽  
Frantz Fanon ◽  
Political Scientist ◽  
Autobiographical Narratives ◽  
Political Ideas ◽  
Dynamic View ◽  
New Generation ◽  
African Intellectuals ◽  
Leopold Sedar Senghor

Esta resenha busca apresentar ao leitor brasileiro a obra Sortir de la grande nuit, do cientista político camaronês Achille Mbembe. Situado na nova geração de pensadores africanos, Achille Mbembe defende uma linha de pensamento a que chama de “afropolitanismo”, a qual busca oferecer uma alternativa aos movimentos da negritude e do panafricanismo. A análise da atual situação cultural e social africana que Mbembe oferece, não obstante seu caráter eminentemente político, utiliza de extenso material autobiográfico, partindo de relatos de suas vivências de juventude em Camarões e incluindo suas experiências como estudante e professor na França, nos Estados Unidos e na África do Sul. Além disso, o autor explicita a influência que tiveram sobre seu pensamento as obras de intelectuais africanos que o precederam, como Frantz Fanon e Leopold Sedar Senghor. Essa miríade de experiências e leituras é harmonizada por Mbembe em uma visão dinâmica da África como local onde diversos povos e culturas transitam, afastando-se, assim, de postulados universalistas e da rigidez das concepções raciais.Palavras-chave: Achille Mbembe. Afropolitanismo. África contemporânea.This review presents to the Brazilian reader Sortir de la grande nuit, a work by the Cameroonian political scientist Achille Mbembe. Belonging to the new generation of African thinkers, Achille Mbembe advocates for a worldview which he calls afropolitanism, offering an alternative to negritude and panafricanism. Mbembe provides an analysis of current social and cultural situation in Africa that takes into account both political ideas and theories and his own autobiographical narratives, from his youth in Cameroon to his experiences as a student and Professor in France, the United States of America, and South Africa. In addition, the author discusses the influence of different African intellectuals, such as Frantz Fanon and Leopold Sedar Senghor, on his own work. These myriad experiences and readings are harmonized into a dynamic view of Africa as a place of transit of diverse peoples and cultures, thus avoiding universalist postulates and rigid racial conceptions.Keywords: Achille Mbembe. Afropolitanism. Contemporary Africa.

scholarly journals Clavulanate inactivation of Staphylococcus aureus β-lactamase

1989 ◽  
Vol 258 (1) ◽  
pp. 205-209 ◽  
Author(s):  
I Rizwi ◽  
A K Tan ◽  
A L Fink ◽  
R Virden
Keyword(s):  
Staphylococcus Aureus ◽  
Catalytic Activity ◽  
Clavulanic Acid ◽  
Low Temperatures ◽  
Stable Form ◽  
Beta Lactamase ◽  
Trans Form ◽  
Conformational Effects ◽  
Enzyme Intermediate ◽  
Enamine Formation

The interaction of clavulanic acid with beta-lactamase from Staphylococcus aureus was investigated, particularly with a view to determining whether conformational effects are involved. The inactivation at neutral pH is essentially stoichiometric, leading to an inactive species with an enamine chromophore. Two forms of the enamine were observed, the first-formed having a positive ellipticity with a maximum near 290 nm. This species slowly converted into the stable form of the inactivated enzyme that had a negative ellipticity with a minimum at 275 nm. This change in sign of the ellipticity of the enamine is consistent with the previously proposed cis-trans isomerization of the enamine [Cartwright & Coulson (1979) Nature (London) 278, 360-361). Both the far-u.v.c.d. and the intrinsic viscosity of the inactivated enzyme indicated that negligible change in conformation of the enzyme accompanied inactivation. The rates of inactivation and enamine formation were compared at low temperatures, where the initial rates were slow enough to be monitored. The rate of loss of 95% of the catalytic activity was almost 100-fold faster than the rate of formation of the first-formed enamine species. The remaining 5% activity was lost with a rate comparable with that for formation of the initial enamine. The simplest explanation of these results is that a relatively stable acyl-enzyme intermediate builds up initially and more slowly partitions between turnover (hydrolysis) and enamine formation. The initially formed enamine is in the cis conformation but slowly isomerizes to the more stable trans form.

scholarly journals Allosteric coupling between α-rings of the 20S proteasome

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Zanlin Yu ◽  
Yadong Yu ◽  
Feng Wang ◽  
Alexander G. Myasnikov ◽  
Philip Coffino ◽  
...  
Keyword(s):  
Protein Degradation ◽  
Cooperative Binding ◽  
20S Proteasome ◽  
Allosteric Coupling ◽  
In Cells

Abstract Proteasomal machinery performs essential regulated protein degradation in eukaryotes. Classic proteasomes are symmetric, with a regulatory ATPase docked at each end of the cylindrical 20S. Asymmetric complexes are also present in cells, either with a single ATPase or with an ATPase and non-ATPase at two opposite ends. The mechanism that populates these different proteasomal complexes is unknown. Using archaea homologs, we construct asymmetric forms of proteasomes. We demonstrate that the gate conformation of the two opposite ends of 20S are coupled: binding one ATPase opens a gate locally, and also opens the opposite gate allosterically. Such allosteric coupling leads to cooperative binding of proteasomal ATPases to 20S and promotes formation of proteasomes symmetrically configured with two identical ATPases. It may also promote formation of asymmetric complexes with an ATPase and a non-ATPase at opposite ends. We propose that in eukaryotes a similar mechanism regulates the composition of the proteasomal population.

scholarly journals Allosteric coupling between α-rings of the 20S proteasome

2019 ◽  
Author(s):  
Zanlin Yu ◽  
Yadong Yu ◽  
Feng Wang ◽  
Alexander G. Myasnikov ◽  
Philip Coffino ◽  
...  
Keyword(s):  
Protein Degradation ◽  
Cooperative Binding ◽  
20S Proteasome ◽  
Allosteric Coupling ◽  
In Cells

AbstractThe proteasomal machinery performs essential regulated protein degradation in eukaryotes. Classic proteasomes are symmetric, with a regulatory ATPase docked at each end of the cylindrical 20S. Asymmetric complexes are also present in cells, either with a single ATPase or with an ATPase and non-ATPase at two opposite ends. The mechanism that populates these different proteasomal complexes is unknown. Using archaea homologs, we constructed asymmetric forms of proteasomes. We demonstrate that the gate conformation of two opposite ends of 20S are coupled: binding one ATPase opens a gate locally, and also the remote opposite gate allosterically. Such allosteric coupling leads to cooperative binding of proteasomal ATPases to 20S, and promotes formation of proteasomes symmetrically configured with two identical ATPases. It may also promote formation of asymmetric complexes with an ATPase and a non-ATPase at opposite ends. We propose that in eukaryotes a similar mechanism regulates the composition of the proteasomal population.

scholarly journals A role for serine-175 in modulating the molecular conformation of calponin

2000 ◽  
Vol 350 (2) ◽  
pp. 579-588 ◽  
Author(s):  
Jian-Ping JIN ◽  
Michael P. WALSH ◽  
Cindy SUTHERLAND ◽  
Wenhua CHEN
Keyword(s):  
Conformational Changes ◽  
Thin Filament ◽  
Molecular Conformation ◽  
Regulatory Function ◽  
Structural Flexibility ◽  
Thin Filaments ◽  
Kinase C ◽  
Conformational Effects ◽  
Myosin Interaction ◽  
Functional States

Calponin is an actin filament-associated protein found in smooth muscle and non-muscle cells. Calponin inhibits actin–myosin interaction in a manner that is prevented by protein kinase C (PKC)-catalysed phosphorylation of serine-175. To investigate the molecular basis of serine-175-mediated regulation, we examined the effect of phosphorylation on the conformation of calponin using monoclonal antibody (mAb) epitope analysis. Eight mAbs against different epitopes on chicken gizzard calponin were developed to monitor the conformational changes in calponin induced by PKC-mediated phosphorylation or serine-175 → alanine (S175A) substitution. The relative affinities of the mAbs for calponins immobilized on microtitre plates or bound to actin–tropomyosin thin filaments were determined, and epitope competitions between free and immobilized calponins were carried out. The changes in binding affinity between mAb paratopes and calponin epitopes demonstrate several serine-175 modification-induced conformational effects: (a) structures of calponin are reconfigured by serine-175 modification, supporting the regulatory function of serine-175; (b) there are submolecular structures unaffected by modification of serine-175 in both free and thin filament-associated calponins, suggesting that the serine-175-based conformational modulation is a targeted allosteric effect; (c) significant conformational changes are detected between free and thin filament-associated calponins, indicating two functional states of the molecular conformation; and (d) the different epitope characteristics between thin filament-bound and free calponins suggest that calponin is a flexible molecule, and the modifications of serine-175 may also determine the structural flexibility to increase the epitope accessibility. These results provide novel information concerning the structure–function relationships of calponin and its regulation by phosphorylation.

scholarly journals Variably modulated gating of the 26S proteasome by ATP and polyubiquitin

2009 ◽  
Vol 421 (3) ◽  
pp. 397-404 ◽  
Author(s):  
Xiaohua Li ◽  
George N. Demartino
Keyword(s):  
Atp Hydrolysis ◽  
26S Proteasome ◽  
20S Proteasome ◽  
Peptide Substrate ◽  
Peptide Hydrolysis ◽  
Atpase Subunit ◽  
Protein Substrates ◽  
State Process ◽  
Unstructured Proteins ◽  
Substrate Hydrolysis

The 26S proteasome is a 2500 kDa protease complex that degrades polyubiquitylated proteins by a mechanism that requires ATP hydrolysis. It also degrades short non-ubiquitylated peptides and certain unstructured proteins by an energy-independent mechanism that requires bound ATP to maintain its component subcomplexes, the 20S proteasome and PA700, in a functionally assembled state. Proteolysis of both types of substrate requires PA700-induced opening of reversible gates at substrate-access pores of the 20S proteasome. In the present study we demonstrate that the rate of peptide substrate hydrolysis, a functional monitor of gate opening, is regulated variably by multiple effectors. ATPγS (adenosine 5′-[γ-thio]triphosphate) and other non-hydrolysable ATP analogues increased peptide substrate hydrolysis by intact 26S proteasomes. Thus nucleotides that maintained 26S proteasome structure, but did not support ATP hydrolysis or the degradation of polyubiquitylated proteins, promoted enhanced rates of peptide hydrolysis. Polyubiquitin and a peptoid that binds selectively to a single ATPase subunit of PA700 also increased rates of peptide hydrolysis but had disparate effects on rates of ATP hydrolysis. The effect of polyubiquitin was specific for ubiquitin–ubiquitin linkages that supported proteolysis of protein substrates. These results indicate that gating of the 26S proteasome is not a simple two-state process but can be variably modulated. Our results suggest that modulated gating of the proteasome may be an important element of the mechanism of proteolysis of polyubiquitylated proteins.

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