Pseudomonas response regulators produced in an E. coli heterologous expression host exhibit host-derived post-translational phosphorylation

2021 ◽  
Author(s):  
Megan E Garber ◽  
Rodrigo Fregoso ◽  
Julie Lake ◽  
Anne Kakouridis ◽  
Aindrila Mukhopadhyay
Keyword(s):  
Pseudomonas Stutzeri ◽  
Acetyl Phosphate ◽  
Post Translational Modification ◽  
Response Regulators ◽  
Phosphorylation State ◽  
E Coli ◽  
Phosphorylated Proteins ◽  
Exogenous Addition ◽  
The Media ◽  
The Impact

In this report, we systematically characterize 32 response regulators (RRs) from a metal tolerant groundwater isolate, Pseudomonas stutzeri RCH2 to assess the impact of host-derived post-translational phosphorylation. As observed by distinct shifted bands in a phos-tag gel, 12 of the 24 detected RRs show homogenous mixtures of phosphorylated proteins or heterogenous mixtures of unphosphorylated and phosphorylated proteins. By evaluating the phosphorylation state of CzcR and CopR II under varying assay parameters, we found that changes to pH and exogenous addition of phospho-donors (e.g. acetyl phosphate) have little to no effect on phosphorylation state. By applying protein production conditions that decrease the pool of intracellular acetyl-phosphate in E. coli, we found a reduction in the phosphorylated population of CopR II when magnesium was added to the media, but observed no change in phosphorylated population when CopR II is expressed in E. coli BL21 (DE3) ∆pta, a mutant with a metabolic disruption to the acetyl-phosphate pathway. Therefore, the specific mechanism of post-translational phosphorylation of RRs in E. coli remains obscure. These findings show the importance of characterizing the phosphorylations state of proteins when heterologously expressed, since their biochemical and physiological properties are dependent on post-translational modification.

scholarly journals The Protein Interactome of Glycolysis in Escherichia coli

Proteomes ◽  
2021 ◽  
Vol 9 (2) ◽  
pp. 16
Author(s):  
Shomeek Chowdhury ◽  
Stephen Hepper ◽  
Mudassir K. Lodi ◽  
Milton H. Saier ◽  
Peter Uetz
Keyword(s):  
Escherichia Coli ◽  
Protein Interactions ◽  
Allosteric Regulation ◽  
Glycolytic Enzymes ◽  
Protein Protein Interactions ◽  
Post Translational Modification ◽  
Interacting Protein ◽  
E Coli ◽  
Protein Interactome ◽  
The Impact

Glycolysis is regulated by numerous mechanisms including allosteric regulation, post-translational modification or protein-protein interactions (PPI). While glycolytic enzymes have been found to interact with hundreds of proteins, the impact of only some of these PPIs on glycolysis is well understood. Here we investigate which of these interactions may affect glycolysis in E. coli and possibly across numerous other bacteria, based on the stoichiometry of interacting protein pairs (from proteomic studies) and their conservation across bacteria. We present a list of 339 protein-protein interactions involving glycolytic enzymes but predict that ~70% of glycolytic interactors are not present in adequate amounts to have a significant impact on glycolysis. Finally, we identify a conserved but uncharacterized subset of interactions that are likely to affect glycolysis and deserve further study.

scholarly journals Identification of novel protein lysine acetyltransferases inEscherichia coli

2018 ◽  
Author(s):  
David G. Christensen ◽  
Jesse G. Meyer ◽  
Jackson T. Baumgartner ◽  
Alexandria K. D’Souza ◽  
William C. Nelson ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Substrate Specificity ◽  
Lysine Acetylation ◽  
Acetyl Phosphate ◽  
Protein Acetylation ◽  
Active Site Residues ◽  
Post Translational Modification ◽  
Post Translational Modifications ◽  
E Coli ◽  
Enzymatic Mechanism

AbstractPost-translational modifications, such as Nε-lysine acetylation, regulate protein function. Nε-lysine acetylation can occur either non-enzymatically or enzymatically. The non-enzymatic mechanism uses acetyl phosphate (AcP) or acetyl coenzyme A (AcCoA) as acetyl donors to modify an Nε-lysine residue of a protein. The enzymatic mechanism uses Nε-lysine acetyltransferases (KATs) to specifically transfer an acetyl group from AcCoA to Nε-lysine residues on proteins. To date, only one KAT (YfiQ, also known as Pka and PatZ) has been identified inE. coli. Here, we demonstrate the existence of 4 additionalE. coliKATs: RimI, YiaC, YjaB, and PhnO. In a genetic background devoid of all known acetylation mechanisms (most notably AcP and YfiQ) and one deacetylase (CobB), overexpression of these putative KATs elicited unique patterns of protein acetylation. We mutated key active site residues and found that most of them eliminated enzymatic acetylation activity. We used mass spectrometry to identify and quantify the specificity of YfiQ and the four novel KATs. Surprisingly, our analysis revealed a high degree of substrate specificity. The overlap between KAT-dependent and AcP-dependent acetylation was extremely limited, supporting the hypothesis that these two acetylation mechanisms play distinct roles in the post-translational modification of bacterial proteins. We further showed that these novel KATs are conserved across broad swaths of bacterial phylogeny. Finally, we determined that one of the novel KATs (YiaC) and the known KAT (YfiQ) can negatively regulate bacterial migration. Together, these results emphasize distinct and specific non-enzymatic and enzymatic protein acetylation mechanisms present in bacteria.ImportanceNε-lysine acetylation is one of the most abundant and important post-translational modifications across all domains of life. One of the best-studied effects of acetylation occurs in eukaryotes, where acetylation of histone tails activates gene transcription. Although bacteria do not have true histones, Nε-lysine acetylation is prevalent; however, the role of these modifications is mostly unknown. We constructed anE. colistrain that lacked both known acetylation mechanisms to identify four new Nε-lysine acetyltransferases (RimI, YiaC, YjaB, and PhnO). We used mass spectrometry to determine the substrate specificity of these acetyltransferases. Structural analysis of selected substrate proteins revealed site-specific preferences for enzymatic acetylation that had little overlap with the preferences of the previously reported acetyl-phosphate non-enzymatic acetylation mechanism. Finally, YiaC and YfiQ appear to regulate flagellar-based motility, a phenotype critical for pathogenesis of many organisms. These acetyltransferases are highly conserved and reveal deeper and more complex roles for bacterial post-translational modification.

The More Control, the Better?

2014 ◽  
Vol 26 (2) ◽  
pp. 81-91 ◽  
Author(s):  
Jeeyun Oh ◽  
Mun-Young Chung ◽  
Sangyong Han
Keyword(s):  
Mixed Design ◽  
Negative Effects ◽  
Story Structure ◽  
Content Type ◽  
High User ◽  
User Control ◽  
Rigorous Research ◽  
Control Versus ◽  
The Media ◽  
The Impact

Despite of the popularity of interactive movie trailers, rigorous research on one of the most apparent features of these interfaces – the level of user control – has been scarce. This study explored the effects of user control on users’ immersion and enjoyment of the movie trailers, moderated by the content type. We conducted a 2 (high user control versus low user control) × 2 (drama film trailer versus documentary film trailer) mixed-design factorial experiment. The results showed that the level of user control over movie trailer interfaces decreased users’ immersion when the trailer had an element of traditional story structure, such as a drama film trailer. Participants in the high user control condition answered that they were less fascinated with, absorbed in, focused on, mentally involved with, and emotionally affected by the movie trailer than participants in the low user control condition only with the drama movie trailer. The negative effects of user control on the level of immersion for the drama trailer translated into users’ enjoyment. The impact of user control over interfaces on immersion and enjoyment varies depending on the nature of the media content, which suggests a possible trade-off between the level of user control and entertainment outcomes.

The impact of news coverage of Russia in the media on export—import activities

2019 ◽  
pp. 30-44
Author(s):  
Elena A. Fedorovau ◽  
Svetlana O. Musienko ◽  
Igor S. Demin ◽  
Fedor Yu. Fedorov ◽  
Dmitriy O. Afanasyev
Keyword(s):  
News Coverage ◽  
The Media ◽  
The Impact

МАРКЕТИНГОВІ АСПЕКТИ ФУНКЦІОНУВАННЯ ЛІТЕРАТУРНИХ КОНКУРСІВ ТА ПРЕМІЙ

2020 ◽  
pp. 90
Author(s):  
O. Bondar
Keyword(s):  
Publishing House ◽  
Target Audience ◽  
Point Of View ◽  
Information Support ◽  
Literary Field ◽  
Non Profit ◽  
Functional Aspects ◽  
The Media ◽  
Literary Prizes ◽  
The Impact

<p><em>In this study, I have collected and summarized the functional aspects of a literary prize, contest, and rating, which indicate their affiliation with the marketing complex of the publishing house for the first time. For this purpose, I have analyzed and summarized the common concepts of the functioning of literary prizes and contests as advertising tools for publishing activity. Because the previous studies are only focused on the fact of the impact of the prize on the promotion of editions but do not explain it, these aspects have been considered and introduced by me from the book production’s point of view. I investigated that the prizes and the contests in the literary field are effective marketing tools, which meet many publisher’s needs at the same time and can be considered a non-profit form of capital. I have reviewed the works of other authors, who accept that the economic success of the book is rising if the author is a winner of the literary prize or contest. I have found out that the book prize activates the demand for the book, and the literary contest is a tool to track the reader’s reaction to a future publication. In this way, literary prizes and contests can be considered as a way of conducting a marketing dialogue with the target audience. I have focused on the information support of literary national and international prizes and contests by the media, which attracts attention to the book and forms the reader’s interest. The literary prizes and contests are also considered as a way of exploring trends and their changes, familiarization the popular genres among the target audience and fixation the current choice of modern readers. Literary prizes and contests motivate the authors to improve their literary excellence, are the source of new authors and works, and assist in increasing sales of books. However, further research is recommended.</em></p><strong><em>Key words:</em></strong><em> book prize, book rating, literary contest, literary prize, functions of the literary prizes.</em>

The Process of Globalization and Its Impact on Media

2016 ◽  
Vol 1 (4) ◽  
pp. 150
Author(s):  
Veton Zejnullahi
Keyword(s):  
New Technology ◽  
World Order ◽  
Modern Society ◽  
Media Access ◽  
Four Corners ◽  
The World ◽  
The Media ◽  
The Republic ◽  
Human Eyes ◽  
The Impact

The process of globalization, which many times is considered as new world order is affecting all spheres of modern society but also the media. In this paper specifically we will see the impact of globalization because we see changing the media access to global problems in general being listed on these processes. We will see that the greatest difficulties will have small media as such because the process is moving in the direction of creating mega media which thanks to new technology are reaching to deliver news and information at the time of their occurrence through choked the small media. So it is fair to conclude that the rapid economic development and especially the technology have made the world seem "too small" to the human eyes, because for real-time we will communicate with the world with the only one Internet connection, and also all the information are take for the development of events in the four corners of the world and direct from the places when the events happen. Even Albanian space has not left out of this process because the media in the Republic of Albania and the Republic of Kosovo are adapted to the new conditions under the influence of the globalization process. This fact is proven powerful through creating new television packages, written the websites and newspapers in their possession.

Impact of Raw Materials and Processing Techniques on The Microbiological Quality of Egyptian Domiati Cheese

2020 ◽  
Keyword(s):  
Raw Materials ◽  
Microbiological Quality ◽  
Raw Milk ◽  
Plate Count ◽  
E Coli ◽  
Fresh Cheese ◽  
Processing Techniques ◽  
The Impact ◽  
Total Yeast

Domiati cheese is the most popular brand of cheese ripened in brine in the Middle East in terms of consumed quantities. This study was performed to investigate the impact of the microbiological quality of the used raw materials, the applied traditional processing techniques and ripening period on the quality and safety of the produced cheese. Three hundred random composite samples were collected from three factories at Fayoum Governorate, Egypt. Collected samples represent twenty-five each of: raw milk, table salt, calf rennet, microbial rennet, water, environmental air, whey, fresh cheese, ripened cheese & swabs from: worker hands; cheese molds and utensils; tanks. All samples were examined microbiologically for Standard Plate Count (SPC), coliforms count, Staphylococcus aureus (S. aureus) count, total yeast & mould count, presence of E. coli, Salmonellae and Listeria monocytogenes (L. monocytogenes). The mean value of SPC, coliforms, S. aureus and total yeast & mould counts ranged from (79×102 CFU/m3 for air to 13×108 CFU/g for fresh cheese), (7×102 MPN/ cm2 for tank swabs to 80×106 MPN/ml for raw milk), (9×102 CFU/g for salt to 69×106 CFU/g for fresh cheese) and (2×102 CFU/cm2 for hand swabs to 60×104 CFU/g for fresh cheese), respectively. Whereas, E. coli, Salmonella and L. monocytogenes failed to be detected in all examined samples. There were significant differences in all determined microbiological parameters (p ≤0.05) between fresh and ripened cheese which may be attributed to different adverse conditions such as water activity, pH, salt content and temperature carried out to improve the quality of the product.

The Health of Refugees

2019 ◽  
Keyword(s):  
Age Groups ◽  
Refugee Health ◽  
Mass Movements ◽  
Ethical Challenges ◽  
Displaced Populations ◽  
Political Landscape ◽  
The Media ◽  
Multidisciplinary Perspective ◽  
The Impact

There have been significant changes in the numbers, patterns, and circumstances of refugees and in the political landscape to support humanitarianism since the publication of the first edition of this collection. Like the first edition, this volume provides a multidisciplinary perspective on refugee health, tracing the health repercussions on individuals and populations from the drivers of forced mass movements of populations from situations of conflict and other disasters through to the process of resettlement in countries other than their countries of origin. Drawing on the expertise of academics, practitioners, and UN frontline experts, the collection covers three main aspects of refugee health: the concepts, definitions, and context from a human rights, humanitarianism, and social determinants of health perspective; the intersection of vulnerabilities across age groups and settings; and the ethical challenges for practitioners and researchers working with forcibly displaced populations seeking to resettle. The collection concludes with an analysis of the role of the media in shaping our perceptions of refugees and the impact on policy and access to care.

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