scholarly journals Abiological Catalysis by Myoglobin Mutant with a Genetically Incorporated Unnatural Amino Acid

2021 ◽  
Author(s):  
Subhash Chand ◽  
Sriparna Ray ◽  
Poonam Yadav ◽  
Susruta Samanta ◽  
Brad S Pierce ◽  
...  
Keyword(s):  
Unnatural Amino Acids ◽  
Oxidation Process ◽  
Reference Electrode ◽  
Unnatural Amino Acid ◽  
Genetically Engineered ◽  
Oxygen Storage ◽  
Compound I ◽  
Redox Active ◽  
Additional Hydrogen

To inculcate biocatalytic activity in the oxygen-storage protein myoglobin (Mb), a genetically engineered myoglobin mutant H64DOPA (DOPA = L-3,4-dihydroxyphenylalanine) has been created.  Incorporation of unnatural amino acids has already demonstrated their ability to accomplish many non-natural functions in proteins efficiently.  Herein, the presence of redox-active DOPA residue in the active site of mutant Mb presumably stabilizes the compound I in the catalytic oxidation process by participating in an additional hydrogen bonding (H-bonding) as compared to the WT Mb.  Specifically, a general acid-base catalytic pathway was achieved due to the availability of the hydroxyl moieties of DOPA.  The reduction potential values of WT (E° = - 260 mV) and mutant Mb (E° = - 300 mV), w.r.t. Ag/AgCl reference electrode, in the presence of hydrogen peroxide, indicated an additional H-bonding in the mutant protein, which is responsible for the peroxidase activity of the mutant Mb.  We observed that in the presence of 5 mM H2O2, H64DOPA Mb oxidizes thioanisole and benzaldehyde with a 10 and 54 folds higher rate, respectively, as opposed to WT Mb.  Based on spectroscopic, kinetic, and electrochemical studies, we deduce that DOPA residue, when present within the distal pocket of mutant Mb, alone serves the role of His/Arg-pair of peroxidases.

Activation of cytochrome c to a peroxidase compound I-type intermediate by H2O2: relevance to redox signalling in apoptosis

2004 ◽  
Vol 71 ◽  
pp. 97-106 ◽  
Author(s):  
Mark Burkitt ◽  
Clare Jones ◽  
Andrew Lawrence ◽  
Peter Wardman
Keyword(s):  
Cytochrome C ◽  
Hydroxyl Radical ◽  
Redox Regulation ◽  
Chemotherapeutic Agents ◽  
Tyrosyl Radical ◽  
Compound I ◽  
Definitive Evidence ◽  
Enhanced Production ◽  
Physico Chemical

The release of cytochrome c from mitochondria during apoptosis results in the enhanced production of superoxide radicals, which are converted to H2O2 by Mn-superoxide dismutase. We have been concerned with the role of cytochrome c/H2O2 in the induction of oxidative stress during apoptosis. Our initial studies showed that cytochrome c is a potent catalyst of 2′,7′-dichlorofluorescin oxidation, thereby explaining the increased rate of production of the fluorophore 2′,7′-dichlorofluorescein in apoptotic cells. Although it has been speculated that the oxidizing species may be a ferryl-haem intermediate, no definitive evidence for the formation of such a species has been reported. Alternatively, it is possible that the hydroxyl radical may be generated, as seen in the reaction of certain iron chelates with H2O2. By examining the effects of radical scavengers on 2′,7′-dichlorofluorescin oxidation by cytochrome c/H2O2, together with complementary EPR studies, we have demonstrated that the hydroxyl radical is not generated. Our findings point, instead, to the formation of a peroxidase compound I species, with one oxidizing equivalent present as an oxo-ferryl haem intermediate and the other as the tyrosyl radical identified by Barr and colleagues [Barr, Gunther, Deterding, Tomer and Mason (1996) J. Biol. Chem. 271, 15498-15503]. Studies with spin traps indicated that the oxo-ferryl haem is the active oxidant. These findings provide a physico-chemical basis for the redox changes that occur during apoptosis. Excessive changes (possibly catalysed by cytochrome c) may have implications for the redox regulation of cell death, including the sensitivity of tumour cells to chemotherapeutic agents.

scholarly journals Tenascin-C in Heart Diseases—The Role of Inflammation

2021 ◽  
Vol 22 (11) ◽  
pp. 5828
Author(s):  
Kyoko Imanaka-Yoshida
Keyword(s):  
Ventricular Remodeling ◽  
Heart Diseases ◽  
Genetically Engineered ◽  
Matricellular Protein ◽  
Myocardial Repair ◽  
Inflammatory Reactions ◽  
Tenascin C ◽  
Genetically Engineered Mice

Tenascin-C (TNC) is a large extracellular matrix (ECM) glycoprotein and an original member of the matricellular protein family. TNC is transiently expressed in the heart during embryonic development, but is rarely detected in normal adults; however, its expression is strongly up-regulated with inflammation. Although neither TNC-knockout nor -overexpressing mice show a distinct phenotype, disease models using genetically engineered mice combined with in vitro experiments have revealed multiple significant roles for TNC in responses to injury and myocardial repair, particularly in the regulation of inflammation. In most cases, TNC appears to deteriorate adverse ventricular remodeling by aggravating inflammation/fibrosis. Furthermore, accumulating clinical evidence has shown that high TNC levels predict adverse ventricular remodeling and a poor prognosis in patients with various heart diseases. Since the importance of inflammation has attracted attention in the pathophysiology of heart diseases, this review will focus on the roles of TNC in various types of inflammatory reactions, such as myocardial infarction, hypertensive fibrosis, myocarditis caused by viral infection or autoimmunity, and dilated cardiomyopathy. The utility of TNC as a biomarker for the stratification of myocardial disease conditions and the selection of appropriate therapies will also be discussed from a clinical viewpoint.

Plant Resistance in Some Modern Soybean Varieties May Favor Population Growth and Modify the Stylet Penetration of Bemisia tabaci (Hemiptera: Aleyrodidae)

2021 ◽  
Author(s):  
Mauricélia F Almeida ◽  
Clébson S Tavares ◽  
Euires O Araújo ◽  
Marcelo C Picanço ◽  
Eugênio E Oliveira ◽  
...  
Keyword(s):  
Population Growth ◽  
Bemisia Tabaci ◽  
Intrinsic Rate ◽  
Reproductive Rate ◽  
Genetically Engineered ◽  
Population Increase ◽  
Net Reproductive Rate ◽  
Bt Toxin ◽  
Demographic Performance

Abstract Complaints of severe damage by whiteflies in soybean fields containing genetically engineered (GE) varieties led us to investigate the role of transgenic soybean varieties expressing resistance to some insects (Cry1Ac Bt toxin) and to herbicide (glyphosate) on the population growth and feeding behavior of Bemisia tabaci (Gennadius) MEAM1 (Hemiptera: Aleyrodidae). In the laboratory, the whiteflies reared on the GE Bt soybeans had a net reproductive rate (R0) 100% higher and intrinsic rate of population increase (rm) 15% higher than those reared on non-GE soybeans. The increased demographic performance was associated with a higher lifetime fecundity. In electrical penetration graphs, the whiteflies reared on the GE soybeans had fewer probes and spent 50% less time before reaching the phloem phase from the beginning of the first successful probe, indicating a higher risk of transmission of whitefly-borne viruses. Data from Neotropical fields showed a higher population density of B. tabaci on two soybean varieties expressing glyphosate resistance and Cry1Ac Bt toxin. These results indicate that some GE soybean varieties expressing insect and herbicide resistances can be more susceptible to whiteflies than non-GE ones or those only expressing herbicide resistance. Most likely, these differences are related to varietal features that increase host-plant susceptibility to whiteflies. Appropriate pest management may be needed to deal with whiteflies in soybean fields, especially in warm regions, and breeders may want to consider the issue when developing new soybean varieties.

scholarly journals Dynamics of Oxidation of Reduced Forms of CO2 under Electrochemical and Open-Сircuit Conditions on Polycrystalline Pt in H2CO3

Metals ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 274
Author(s):  
Alexander V. Smolin ◽  
Мikhail N. Mikhailov ◽  
Aleksey F. Gadzaov ◽  
Leonid M. Kustov
Keyword(s):  
Quantum Chemical ◽  
Preliminary Analysis ◽  
Oxidation Reaction ◽  
Density Functional ◽  
Oxidation Process ◽  
Open Circuit ◽  
Organic Substances ◽  
Oxidation Of Co ◽  
Reduced Forms

The problem of identifying correlations between catalytic and electrocatalytic processes is one of the fundamental problems of catalysis among “simple” organic substances, and the oxidation of CO and rCO2 is of great interest, since CO and CO2 are considered in pairs both during catalytic and electrocatalytic transformations. In the case of electrocatalysis, this analysis is important in the study of fuel cells. In this paper, we studied the correlation between the oxidation of reduced forms of CO2 (rCO2) under potentiodynamic-galvanoctatic electrochemical and open-circuit conditions of measurements on polycrystalline (pc)Pt in H2CO3. Periodic oscillations are revealed at the oxidation of Had and rCO2 on (pc)Pt. Quantum chemical calculations were carried out on the Pt13 cluster in order to identify the mechanisms of the rCO2 oxidation reaction. The correspondence in the energy parameters of the oxidation process of rCO2 under open-circuit conditions and electrochemical conditions is shown. The preliminary analysis of the system using density functional (DFT) calculations is carried out and the most stable systems that are based on Pt13 are found, namely rOH-Pt13-(CO)n, rOH-Pt13-(COH) and rOH-Pt13-(rCOOH). OH• species was chosen as the most likely candidate for the role of the oxidant for rCO2. Preliminary calculations for the expected reactions were carried out, and the optimal PES is revealed.

scholarly journals Transferability of N-terminal mutations of pyrrolysyl-tRNA synthetase in one species to that in another species on unnatural amino acid incorporation efficiency

Amino Acids ◽  
2020 ◽  
Author(s):  
Thomas L. Williams ◽  
Debra J. Iskandar ◽  
Alexander R. Nödling ◽  
Yurong Tan ◽  
Louis Y. P. Luk ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Genetic Code ◽  
Unnatural Amino Acids ◽  
Trna Synthetase ◽  
Unnatural Amino Acid ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Genetic Code Expansion ◽  
Incorporation Efficiency

AbstractGenetic code expansion is a powerful technique for site-specific incorporation of an unnatural amino acid into a protein of interest. This technique relies on an orthogonal aminoacyl-tRNA synthetase/tRNA pair and has enabled incorporation of over 100 different unnatural amino acids into ribosomally synthesized proteins in cells. Pyrrolysyl-tRNA synthetase (PylRS) and its cognate tRNA from Methanosarcina species are arguably the most widely used orthogonal pair. Here, we investigated whether beneficial effect in unnatural amino acid incorporation caused by N-terminal mutations in PylRS of one species is transferable to PylRS of another species. It was shown that conserved mutations on the N-terminal domain of MmPylRS improved the unnatural amino acid incorporation efficiency up to five folds. As MbPylRS shares high sequence identity to MmPylRS, and the two homologs are often used interchangeably, we examined incorporation of five unnatural amino acids by four MbPylRS variants at two temperatures. Our results indicate that the beneficial N-terminal mutations in MmPylRS did not improve unnatural amino acid incorporation efficiency by MbPylRS. Knowledge from this work contributes to our understanding of PylRS homologs which are needed to improve the technique of genetic code expansion in the future.

scholarly journals Targeting HSPA1A in ARID2-deficient lung adenocarcinoma

2021 ◽  
Author(s):  
Xue Wang ◽  
Yuetong Wang ◽  
Zhaoyuan Fang ◽  
Hua Wang ◽  
Jian Zhang ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Lung Adenocarcinoma ◽  
Genetically Engineered ◽  
Malignant Progression ◽  
Murine Models ◽  
Rna Seq ◽  
Potential Therapeutic Target ◽  
Lung Cancers ◽  
Integrative Analyses

Abstract Somatic mutations of the chromatin remodeling gene ARID2 are observed in about 7% of human lung adenocarcinoma (LUAD). However, the role of ARID2 in the pathogenesis of LUAD remains largely unknown. Here we find that ARID2 expression is decreased during the malignant progression of both human and mice LUAD. Using two KrasG12D-based genetically engineered murine models (GEMM), we demonstrate that ARID2 knockout significantly promotes lung cancer malignant progression and shortens the overall survival. Consistently, ARID2 knockdown significantly promotes cell proliferation in human and mice lung cancer cells. Through integrative analyses of Chip-Seq and RNA-Seq data, we find that Hspa1a is up-regulated by Arid2 loss. Knockdown of Hspa1a specifically inhibits malignant progression of Arid2-deficient but not Arid2-wt lung cancers in both cell lines as well as animal models. Treatment with Hspa1a inhibitor could significantly inhibit the malignant progression of lung cancer with Arid2 deficiency. Together, our findings establish ARID2 as an important tumor suppressor in LUAD with novel mechanistic insights, and further identify HSPA1A as a potential therapeutic target in ARID2-deficient LUAD.

Oxygen and CO Adsorption on Au/SiO2Catalysts Prepared by Magnetron Sputtering: The Role of Oxygen Storage

2010 ◽  
Vol 49 (21) ◽  
pp. 10428-10437 ◽  
Author(s):  
Xiaolin Zheng ◽  
Gabriel M. Veith ◽  
Evgeniy Redekop ◽  
Cynthia S. Lo ◽  
Gregory S. Yablonsky ◽  
...  
Keyword(s):  
Magnetron Sputtering ◽  
Co Adsorption ◽  
Oxygen Storage

scholarly journals miR-146a is a significant brake on autoimmunity, myeloproliferation, and cancer in mice

2011 ◽  
Vol 208 (6) ◽  
pp. 1189-1201 ◽  
Author(s):  
Mark P. Boldin ◽  
Konstantin D. Taganov ◽  
Dinesh S. Rao ◽  
Lili Yang ◽  
Jimmy L. Zhao ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Immune Cell ◽  
Myeloid Cell ◽  
Genetically Engineered ◽  
Biological Processes ◽  
Targeted Deletion ◽  
Genetically Engineered Mice ◽  
Immune Defects ◽  
Molecular Brake

Excessive or inappropriate activation of the immune system can be deleterious to the organism, warranting multiple molecular mechanisms to control and properly terminate immune responses. MicroRNAs (miRNAs), ∼22-nt-long noncoding RNAs, have recently emerged as key posttranscriptional regulators, controlling diverse biological processes, including responses to non-self. In this study, we examine the biological role of miR-146a using genetically engineered mice and show that targeted deletion of this gene, whose expression is strongly up-regulated after immune cell maturation and/or activation, results in several immune defects. Collectively, our findings suggest that miR-146a plays a key role as a molecular brake on inflammation, myeloid cell proliferation, and oncogenic transformation.

Computational Study on Compound I Redox-Active Species in Horseradish Peroxydase Enzyme: Conformational Fluctuations and Solvation Effects

2010 ◽  
Vol 114 (20) ◽  
pp. 6817-6824 ◽  
Author(s):  
Costantino Zazza ◽  
Amedeo Palma ◽  
Nico Sanna ◽  
Simone Tatoli ◽  
Massimiliano Aschi
Keyword(s):  
Computational Study ◽  
Active Species ◽  
Compound I ◽  
Solvation Effects ◽  
Redox Active
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