pH-Dependent Structural Changes in the Active Site ofp-Hydroxybenzoate Hydroxylase Point to the Importance of Proton and Water Movements during Catalysis†,‡

Biochemistry ◽  
1996 ◽  
Vol 35 (2) ◽  
pp. 567-578 ◽  
Author(s):  
Domenico L. Gatti ◽  
Barrie Entsch ◽  
David P. Ballou ◽  
Martha L. Ludwig
Keyword(s):  
Active Site ◽  
Structural Changes ◽  
Ph Dependent ◽  
Hydroxybenzoate Hydroxylase

scholarly journals Cofilin is a pH sensor for actin free barbed end formation: role of phosphoinositide binding

2008 ◽  
Vol 183 (5) ◽  
pp. 865-879 ◽  
Author(s):  
Christian Frantz ◽  
Gabriela Barreiro ◽  
Laura Dominguez ◽  
Xiaoming Chen ◽  
Robert Eddy ◽  
...  
Keyword(s):  
Actin Filament ◽  
Structural Changes ◽  
Ph Sensor ◽  
Ph Sensitive ◽  
Actin Binding ◽  
Filamentous Actin ◽  
Filament Dynamics ◽  
Ph Dependent ◽  
Dynamics Simulations

Newly generated actin free barbed ends at the front of motile cells provide sites for actin filament assembly driving membrane protrusion. Growth factors induce a rapid biphasic increase in actin free barbed ends, and we found both phases absent in fibroblasts lacking H+ efflux by the Na-H exchanger NHE1. The first phase is restored by expression of mutant cofilin-H133A but not unphosphorylated cofilin-S3A. Constant pH molecular dynamics simulations and nuclear magnetic resonance (NMR) reveal pH-sensitive structural changes in the cofilin C-terminal filamentous actin binding site dependent on His133. However, cofilin-H133A retains pH-sensitive changes in NMR spectra and severing activity in vitro, which suggests that it has a more complex behavior in cells. Cofilin activity is inhibited by phosphoinositide binding, and we found that phosphoinositide binding is pH-dependent for wild-type cofilin, with decreased binding at a higher pH. In contrast, phosphoinositide binding by cofilin-H133A is attenuated and pH insensitive. These data suggest a molecular mechanism whereby cofilin acts as a pH sensor to mediate a pH-dependent actin filament dynamics.

scholarly journals The Loop of the TPR1 Subdomain of Phi29 DNA Polymerase Plays a Pivotal Role in Primer-Terminus Stabilization at the Polymerization Active Site

Biomolecules ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 648
Author(s):  
del Prado ◽  
Santos ◽  
Lázaro ◽  
Salas ◽  
de Vega
Keyword(s):  
Dna Polymerase ◽  
Active Site ◽  
Structural Changes ◽  
Binding Capacity ◽  
Dna Polymerases ◽  
Crystallographic Structure ◽  
Genome Replication ◽  
Terminal Protein ◽  
Phi29 Dna Polymerase

Bacteriophage Phi29 DNA polymerase belongs to the protein-primed subgroup of family B DNA polymerases that use a terminal protein (TP) as a primer to initiate genome replication. The resolution of the crystallographic structure showed that it consists of an N-terminal domain with the exonuclease activity and a C-terminal polymerization domain. It also has two subdomains specific of the protein-primed DNA polymerases; the TP Regions 1 (TPR1) that interacts with TP and DNA, and 2 (TPR2), that couples both processivity and strand displacement to the enzyme. The superimposition of the structures of the apo polymerase and the polymerase in the polymerase/TP heterodimer shows that the structural changes are restricted almost to the TPR1 loop (residues 304–314). In order to study the role of this loop in binding the DNA and the TP, we changed the residues Arg306, Arg308, Phe309, Tyr310, and Lys311 into alanine, and also made the deletion mutant Δ6 lacking residues Arg306–Lys311. The results show a defective TP binding capacity in mutants R306A, F309A, Y310A, and Δ6. The additional impaired primer-terminus stabilization at the polymerization active site in mutants Y310A and Δ6 allows us to propose a role for the Phi29 DNA polymerase TPR1 loop in the proper positioning of the DNA and TP-priming 3’-OH termini at the preinsertion site of the polymerase to enable efficient initiation and further elongation steps during Phi29 TP-DNA replication.

scholarly journals Cheminformatics Modeling of Closantel Analogues for Treating River Blindness

2020 ◽  
Author(s):  
Melaine A. Kuenemann ◽  
Phyo Phyo Zin ◽  
Sravya Kuchibhotla ◽  
Denis Fourches
Keyword(s):  
Active Site ◽  
Structural Changes ◽  
Parasitic Nematode ◽  
Chemical Space ◽  
Therapeutic Agents ◽  
Binding Modes ◽  
River Blindness ◽  
Structure Activity ◽  
Potential Binding ◽  
Biological Target

<p></p><p>Onchocerciasis (also known as river blindness<i>)</i> is a neglected tropical disease caused by the <i>Onchocerca volvulus</i> parasitic nematode. Currently, the only approved drug for treating this disease is ivermectin, which is a broad-spectrum antiparasitic agent. However, signs of resistance towards ivermectin have started to emerge. New therapeutic agents are thus urgently needed. The OvCHT1 chitinase enzyme from <i>O. volvulus</i> has been established as a relevant biological target for combatting river blindness. The veterinary anthelmintic drug closantel has been found to be a potent, micro-molar OvCHT1 inhibitor. Herein, we investigated the chemical space of closantel and all its synthesized analogues, focusing on the analysis of their potential binding modes towards OvCHT1. First, we conducted an unsupervised hierarchical clustering to group highly similar analogues and explore structure-activity relationships. Second, we conducted a structure-based molecular docking to predict and study the binding modes of all 57 closantel analogues in the active site of OvCHT1. Third, we screened more than 4 million lead-like compounds from the ZINC library to identify other structurally similar ligands that could potentially bind to OvCHT1. The cheminformatics analysis of the closantel analogues illustrated how minor structural changes in closantel analogues can impact their OvCHT1 activity.</p><p></p>

P219L substitution in human D-amino acid oxidase impacts the ligand binding and catalytic efficiency

2020 ◽  
Vol 168 (5) ◽  
pp. 557-567
Author(s):  
Wanitcha Rachadech ◽  
Yusuke Kato ◽  
Rabab M Abou El-Magd ◽  
Yuji Shishido ◽  
Soo Hyeon Kim ◽  
...  
Keyword(s):  
Amino Acid ◽  
Conformational Changes ◽  
Active Site ◽  
Structural Changes ◽  
Catalytic Efficiency ◽  
Acid Oxidase ◽  
Wild Type ◽  
Amino Acid Oxidase ◽  
Adenine Ring ◽  
The Impact

Abstract Human D-amino acid oxidase (DAO) is a flavoenzyme that is implicated in neurodegenerative diseases. We investigated the impact of replacement of proline with leucine at Position 219 (P219L) in the active site lid of human DAO on the structural and enzymatic properties, because porcine DAO contains leucine at the corresponding position. The turnover numbers (kcat) of P219L were unchanged, but its Km values decreased compared with wild-type, leading to an increase in the catalytic efficiency (kcat/Km). Moreover, benzoate inhibits P219L with lower Ki value (0.7–0.9 µM) compared with wild-type (1.2–2.0 µM). Crystal structure of P219L in complex with flavin adenine dinucleotide (FAD) and benzoate at 2.25 Å resolution displayed conformational changes of the active site and lid. The distances between the H-bond-forming atoms of arginine 283 and benzoate and the relative position between the aromatic rings of tyrosine 224 and benzoate were changed in the P219L complex. Taken together, the P219L substitution leads to an increase in the catalytic efficiency and binding affinity for substrates/inhibitors due to these structural changes. Furthermore, an acetic acid was located near the adenine ring of FAD in the P219L complex. This study provides new insights into the structure–function relationship of human DAO.

Molecular studies on structural changes and oligomerisation of violaxanthin de-epoxidase associated with the pH-dependent activation

2016 ◽  
Vol 129 (1) ◽  
pp. 29-41 ◽  
Author(s):  
Erik Ingmar Hallin ◽  
Mahmudul Hasan ◽  
Kuo Guo ◽  
Hans-Erik Åkerlund
Keyword(s):  
Structural Changes ◽  
Molecular Studies ◽  
Ph Dependent
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