scholarly journals An evolutionary non-conserved motif in Helicobacter pylori arginase mediates positioning of the loop containing the catalytic residue for catalysis

2021 ◽  
Vol 478 (4) ◽  
pp. 871-894
Author(s):  
Ankita Dutta ◽  
Ditsa Sarkar ◽  
Pooja Murarka ◽  
Tasneem Kausar ◽  
Satya Narayan ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Structural Integrity ◽  
Double Mutation ◽  
Conserved Motif ◽  
Catalytic Function ◽  
Underlying Basis ◽  
Critical Residues ◽  
Dynamics Simulations ◽  
The Individual

The binuclear metalloenzyme Helicobacter pylori arginase is important for pathogenesis of the bacterium in the human stomach. Despite conservation of the catalytic residues, this single Trp enzyme has an insertion sequence (–153ESEEKAWQKLCSL165–) that is extremely crucial to function. This sequence contains the critical residues, which are conserved in the homolog of other Helicobacter gastric pathogens. However, the underlying basis for the role of this motif in catalytic function is not completely understood. Here, we used biochemical, biophysical and molecular dynamics simulations studies to determine that Glu155 of this stretch interacts with both Lys57 and Ser152. These interactions are essential for positioning of the motif through Trp159, which is located near Glu155 (His122–Trp159–Tyr125 contact is essential to tertiary structural integrity). The individual or double mutation of Lys57 and Ser152 to Ala considerably reduces catalytic activity with Lys57 to Ala being more significant, indicating they are crucial to function. Our data suggest that the Lys57–Glu155–Ser152 interaction influences the positioning of the loop containing the catalytic His133 so that this His can participate in catalysis, thereby providing a mechanistic understanding into the role of this motif in catalytic function. Lys57 was also found only in the arginases of other Helicobacter gastric pathogens. Based on the non-conserved motif, we found a new molecule, which specifically inhibits this enzyme. Thus, the present study not only provides a molecular basis into the role of this motif in function, but also offers an opportunity for the design of inhibitors with greater efficacy.

Characterization of Thin Organic and Polymeric Films by Spectroscopic Methods

MRS Bulletin ◽  
1987 ◽  
Vol 12 (8) ◽  
pp. 39-41 ◽  
Author(s):  
J.F. Rabolt
Keyword(s):  
Thin Films ◽  
Structural Integrity ◽  
Materials Science ◽  
Molecular Level ◽  
Polymeric Films ◽  
Organic Films ◽  
Semiconductor Films ◽  
Intimate Contact ◽  
The Individual

Much of the anisotropic mechanical and thermal behavior exhibited by materials can be attributed to anisotropic orientation at the molecular level. In self-supporting thin films (5–10 microns) and those (0.01–1.0 microns) deposited on solid substrates, the role of molecular orientation is even more important since it will critically determine their two-dimensional behavior and their structural integrity as well. These two aspects are extremely important because if thin organic and polymeric films are to be competitive with existing materials for such diverse applications1 as chemical sensors and integrated optics, they must be mechanically robust and, ideally, defect free. These stringent constraints dictate that sophisticated characterization techniques, which can interrogate at the molecular level, be developed or refined so as to have the sensitivity to address these critical issues.The development of nondestructive techniques for studying thin organic films has certainly lagged behind those developed for metallic and semiconductor films. Unfortunately, many of these same techniques cannot be simply applied to organic films because they are “invasive” and often alter the structure of the system they were designed to probe. This is especially so in organic and polymer films, and this awareness within the materials science community has led to the adaptation of many photon intensive techniques to the study of thin films. A number of these will be discussed in later sections with their relative merit put in perspective.Certainly the origins of anisotropic structure in bulk materials are manyfold, but the spatial constraints in 2-D can lead to even more complex causes of orientation. In thin films on substrates the role of the surface is important in determining the ordering and orientation of the individual molecular segments which come into intimate contact with it. The extent of this orientation and order is still somewhat controversial but there is general agreement that it probably differs depending on the nature of the substrate.

scholarly journals ParC and GyrA May Be Interchangeable Initial Targets of Some Fluoroquinolones in Streptococcus pneumoniae

1999 ◽  
Vol 43 (2) ◽  
pp. 302-306 ◽  
Author(s):  
Emmanuelle Varon ◽  
Claire Janoir ◽  
Marie-Dominique Kitzis ◽  
Laurent Gutmann
Keyword(s):  
Streptococcus Pneumoniae ◽  
Susceptible Strain ◽  
Quinolone Resistance ◽  
Topoisomerase Iv ◽  
Double Mutation ◽  
Active Efflux ◽  
Single Target ◽  
Resistant Mutants ◽  
The Individual

ABSTRACT To evaluate the role of known topoisomerase IV and gyrase mutations in the fluoroquinolone (FQ) resistance of Streptococcus pneumoniae, we transformed susceptible strain R6 with PCR-generated fragments encompassing the quinolone resistance-determining regions (QRDRs) of parC orgyrA from different recently characterized FQ-resistant mutants. Considering the MICs of FQs and the GyrA and/or ParC mutations of the individual transformants, we found three levels of resistance. The first level was obtained when a single target, ParC or GyrA, depending on the FQ, was modified. An additional mutation(s) in a second target, GyrA or ParC, led to the second level. The highest increases in resistance levels were seen for Bay y3118 and moxifloxacin with the transformant harboring a double mutation in both ParC and GyrA. When a single modified target was considered, only the ParC mutation(s) led to an increase in the MICs of pefloxacin and trovafloxacin. In contrast, the GyrA or ParC mutation(s) could lead to increases in the MICs of ciprofloxacin, sparfloxacin, grepafloxacin, Bay y3118, and moxifloxacin. These results suggest that the preferential target of trovafloxacin and pefloxacin is ParC, whereas either ParC or GyrA may both be initial targets for the remaining FQs tested. The contribution of the ParC and GyrA mutations to efflux-mediated FQ resistance was also examined. Active efflux was responsible for two- to fourfold increases in the MICs of ciprofloxacin for the transformants, regardless of the initial FQ resistance levels of the recipients.

scholarly journals ptFVa ( Pseudonaja Textilis Venom-Derived Factor Va) Retains Structural Integrity Following Proteolysis by Activated Protein C

2021 ◽  
Author(s):  
Mark Schreuder ◽  
Xiaosong Liu ◽  
Ka Lei Cheung ◽  
Pieter H. Reitsma ◽  
Gerry A.F. Nicolaes ◽  
...  
Keyword(s):  
Protein C ◽  
Structural Integrity ◽  
Noncovalent Interactions ◽  
Activated Protein C ◽  
Regulatory Mechanisms ◽  
Factor V ◽  
Factor Va ◽  
Dynamics Simulations ◽  
A2 Domain

Objective: The Australian snake ptFV ( Pseudonaja textilis venom-derived factor V) variant that retains cofactor function despite APC (activated protein C)-dependent proteolysis. Here, we aimed to unravel the mechanistic principles by determining the role of the absent Arg306 cleavage site that is required for the inactivation of Fva (mammalian factor Va). Approach and Results: Our findings show that in contrast to human FVa, APC-catalyzed proteolysis of ptFVa at Arg306 and Lys507 does not abrogate ptFVa cofactor function. Remarkably, the structural integrity of APC-proteolyzed ptFVa is maintained indicating that stable noncovalent interactions prevent A2-domain dissociation. Using Molecular Dynamics simulations, we uncovered key regions located in the A1 and A2 domain that may be at the basis of this remarkable characteristic. Conclusions: Taken together, we report a completely novel role for uniquely adapted regions in ptFVa that prevent A2 domain dissociation. As such, these results challenge our current understanding by which strict regulatory mechanisms control FVa activity.

scholarly journals A Conserved Structural Role for the Walker-A Lysine in P-Loop Containing Kinases

2021 ◽  
Vol 8 ◽  
Author(s):  
Fatlum Hajredini ◽  
Ranajeet Ghose
Keyword(s):  
Tyrosine Kinases ◽  
Specific Interaction ◽  
Enhanced Sampling ◽  
Structural Role ◽  
Catalytic Function ◽  
Key Characteristics ◽  
Optimal Coordination ◽  
Enzyme Families ◽  
Dynamics Simulations

Bacterial tyrosine kinases (BY-kinases) and shikimate kinases (SKs) comprise two structurally divergent P-loop containing enzyme families that share similar catalytic site geometries, most notably with respect to their Walker-A, Walker-B, and DxD motifs. We had previously demonstrated that in BY-kinases, a specific interaction between the Walker-A and Walker-B motifs, driven by the conserved “catalytic” lysine housed on the former, leads to a conformation that is unable to efficiently coordinate Mg2+•ATP and is therefore incapable of chemistry. Here, using enhanced sampling molecular dynamics simulations, we demonstrate that structurally similar interactions between the Walker-A and Walker-B motifs, also mediated by the catalytic lysine, stabilize a state in SKs that deviates significantly from one that is necessary for the optimal coordination of Mg2+•ATP. This structural role of the Walker-A lysine is a general feature in SKs and is found to be present in members that encode a Walker-B sequence characteristic of the family (Coxiella burnetii SK), and in those that do not (Mycobacterium tuberculosis SK). Thus, the structural role of the Walker-A lysine in stabilizing an inactive state, distinct from its catalytic function, is conserved between two distantly related P-loop containing kinase families, the SKs and the BY-kinases. The universal conservation of this element, and of the key characteristics of its associated interaction partners within the Walker motifs of P-loop containing enzymes, suggests that this structural role of the Walker-A lysine is perhaps a widely deployed regulatory mechanism within this ancient family.

Metal-induced change in catalytic loop positioning in Helicobacter pylori arginase alters catalytic function

2019 ◽  
Vol 476 (23) ◽  
pp. 3595-3614 ◽  
Author(s):  
Ankita Dutta ◽  
Mohit Mazumder ◽  
Mashkoor Alam ◽  
Samudrala Gourinath ◽  
Apurba Kumar Sau
Keyword(s):  
Helicobacter Pylori ◽  
Active Site ◽  
Underlying Mechanism ◽  
Essential Step ◽  
Catalytic Function ◽  
Km Value ◽  
Catalytic Loop ◽  
The Difference ◽  
H Pylori ◽  
Dynamics Simulations

Arginase is a bimetallic enzyme that utilizes mainly Mn2+ or Co2+ for catalytic function. In human homolog, the substitution of Mn2+ with Co2+ significantly reduces the Km value without affecting the kcat. However, in the Helicobacter pylori counterpart (important for pathogenesis), the kcat increases nearly 4-fold with Co2+ ions both in the recombinant holoenzyme and arginase isolated from H. pylori grown with Co2+ or Mn2+. This suggests that the active site of arginase in the two homologs is modulated differently by these two metal ions. To investigate the underlying mechanism for metal-induced difference in catalytic activity in the H. pylori enzyme, we used biochemical, biophysical and microsecond molecular dynamics simulations studies. The study shows that the difference in binding affinity of Co2+ and Mn2+ ions with the protein is linked to a different positioning of a loop (–122HTAYDSDSKHIHG134–) that contains a conserved catalytic His133. Consequently, the proximity of His133 and conserved Glu281 is varied. We found that the Glu281–His133 interaction is crucial for catalytic function and was previously unexplored in other homologs. We suggest that the proximity difference between these two residues in the Co2+- and Mn2+-proteins alters the proportion of protonated His133 via variation in its pKa. This affects the efficiency of proton transfer — an essential step of l-arginine hydrolysis reaction catalyzed by arginase and thus activity. Unlike in human arginase, the flexibility of the above segment observed in H. pylori homolog suggests that this region in the H. pylori enzyme may be explored to design its specific inhibitors.

Peripheral insertion modulates the editing activity of the isolated CP1 domain of leucyl-tRNA synthetase

2011 ◽  
Vol 440 (2) ◽  
pp. 217-227 ◽  
Author(s):  
Ru-Juan Liu ◽  
Min Tan ◽  
Dao-Hai Du ◽  
Bei-Si Xu ◽  
Gilbert Eriani ◽  
...  
Keyword(s):  
Active Site ◽  
Trna Synthetase ◽  
Site Directed Mutagenesis ◽  
Aquifex Aeolicus ◽  
Trna Synthetases ◽  
Catalytic Function ◽  
Editing Activity ◽  
Domain Docking ◽  
Dynamics Simulations

A large insertion domain called CP1 (connective peptide 1) present in class Ia aminoacyl-tRNA synthetases is responsible for post-transfer editing. LeuRS (leucyl-tRNA synthetase) from Aquifex aeolicus and Giardia lamblia possess unique 20 and 59 amino acid insertions respectively within the CP1 that are crucial for editing activity. Crystal structures of AaLeuRS-CP1 [2.4 Å (1 Å=0.1 nm)], GlLeuRS-CP1 (2.6 Å) and the insertion deletion mutant AaLeuRS-CP1Δ20 (2.5 Å) were solved to understand the role of these insertions in editing. Both insertions are folded as peripheral motifs located on the opposite side of the proteins from the active-site entrance in the CP1 domain. Docking modelling and site-directed mutagenesis showed that the insertions do not interact with the substrates. Results of molecular dynamics simulations show that the intact CP1 is more dynamic than its mutant devoid of the insertion motif. Taken together, the data show that a peripheral insertion without a substrate-binding site or major structural role in the active site may modulate catalytic function of a protein, probably from protein dynamics regulation in two respective LeuRS CP1s. Further results from proline and glycine mutational analyses intended to reduce or increase protein flexibility are consistent with this hypothesis.

School Success and School Engagement of Immigrant Children and Adolescents

2013 ◽  
Vol 18 (2) ◽  
pp. 126-135 ◽  
Author(s):  
Frosso Motti-Stefanidi ◽  
Ann S. Masten
Keyword(s):  
Academic Achievement ◽  
Academic Success ◽  
Children And Adolescents ◽  
School Engagement ◽  
School Success ◽  
Immigrant Children ◽  
Integrative Model ◽  
High Stakes ◽  
The Individual

Academic achievement in immigrant children and adolescents is an indicator of current and future adaptive success. Since the future of immigrant youths is inextricably linked to that of the receiving society, the success of their trajectory through school becomes a high stakes issue both for the individual and society. The present article focuses on school success in immigrant children and adolescents, and the role of school engagement in accounting for individual and group differences in academic achievement from the perspective of a multilevel integrative model of immigrant youths’ adaptation ( Motti-Stefanidi, Berry, Chryssochoou, Sam, & Phinney, 2012 ). Drawing on this conceptual framework, school success is examined in developmental and acculturative context, taking into account multiple levels of analysis. Findings suggest that for both immigrant and nonimmigrant youths the relationship between school engagement and school success is bidirectional, each influencing over time the other. Evidence regarding potential moderating and mediating roles of school engagement for the academic success of immigrant youths also is evaluated.

Sign in / Sign up

Export Citation Format

Share Document