Download or read book Characterization of the Nitric Oxide Sensing Domain of Soluable Guanylate Cyclase written by Yunde Zhao and published by . This book was released on 1999 with total page 474 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Identification and Characterization of a Conserved Soluble Guanylate Cyclase like Heme Domain that Discriminates Between Nitric Oxide and Oxygen written by David Stanley Karow and published by . This book was released on 2004 with total page 298 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book cGMP Generators Effectors and Therapeutic Implications written by Harald H. H. W. Schmidt and published by Springer Science & Business Media. This book was released on 2008-12-17 with total page 581 pages. Available in PDF, EPUB and Kindle. Book excerpt: After the discovery of endogenous NO formation in the late '80s and the 1998 Nobel Prize in Physiology or Medicine, many researchers and physicians again became interested in the NO/sGC interaction and cGMP-dependent signaling. This book is an enthusiastic celebration of cyclic guanosine monophosphate (cGMP) and amply illustrates the importance of this field of science to patients and the way in which the field has evolved. It is exclusively devoted to this exciting and important signaling molecule, addressing all recent advances in understanding guanylate cyclase regulation, NO/sGC interactions, cGMP effector mechanisms and their pathophysiological and pharmacological implications. Particular attention will also be given to clinical applications of the novel cGMP-elevating drugs which are on the horizon, thus spanning the continuum from basic science to clinic.

Download or read book Functional and Mechanistic Characterization of Bacterial Nitric Oxide Signaling Pathways written by Minxi Rao and published by . This book was released on 2016 with total page 96 pages. Available in PDF, EPUB and Kindle. Book excerpt: Nitric oxide (NO) is a well-established signaling molecule and cytotoxic agent in mammals. NO is synthesized by nitric oxide synthase (NOS) by macrophages at high concentrations as a key part of the host immune response, and at low concentrations in endothelial and neuronal cells as a signaling agent. In endothelial cells, the primary NO receptor is soluble guanylate cyclase (sGC), which contains a heme-nitric oxide/oxygen binding domain (H-NOX). Selective binding of NO to the H-NOX domain is responsible for activation of sGC. Thus, the mammalian NO signaling system involves NO synthesis by NOS, and NO sensing by the H-NOX domain of sGC. NOS and H-NOX proteins have also been identified in a number of bacterial species, including pathogens. Putative roles for bacterial NOS proteins include protection against oxidative stress and antibiotics, while bacterial H-NOX proteins have been shown to govern processes such as biofilm formation and bioluminescence via interactions with signaling proteins such as diguanylate cyclases (DGC) or histidine kinases (HK). Here, various aspects of NO signaling from three different organisms are characterized: the marine alphaproteobacterium Silicibacter sp. TrichCH4B; the soil-dwelling gammaproteobacterium Shewanella oneidensis, and the marine cyanobacterium Synechococcus sp. PCC 7335. This work and other recent studies seek to understand not only the diverse roles for NO in bacteria, but also the molecular mechanisms of bacterial NO signaling. Silicibacter sp. TrichCH4B is the first bacterial organism discovered to contain both an NOS and H-NOX, thus capable of both NO synthesis and sensing, analogous to mammalian systems. The H-NOX protein from Silicibacter is found in an operon adjacent to an HK, forming part of a two-component phospho-relay signaling network. The response regulator of the network was identified to be a diguanylate cyclase (DGC), which is inactivated upon phosphorylation and establishes the link between NO and intracellular cyclic-di-GMP levels, and consequently biofilm formation. It was also determined that Silicibacter NOS activity is stimulated by a signaling protein from an algal symbiont, Trichodesmium erythraeum, which is a major marine nitrogen fixer. Thus, in the presence of Trichodesmium, the increase in NOS activity results in Silicibacter biofilm formation and poising the two species for nutrient exchange, revealing a novel role for NO in interspecies communication and symbiosis. Given the diverse processes governed by NO/H-NOX signaling, it is crucial to understand the molecular mechanism by which H-NOX regulates HK autophosphorylation activity, the most common outcome of a NO-bound H-NOX. Here, the interaction and signal transduction between the H-NOX-HK signaling pair from Shewanella oneidensis are characterized. Binding kinetics measurements and analytical gel filtration revealed that NO-bound H-NOX has a tighter affinity for the HK, compared with H-NOX in the unliganded state, correlating binding affinity with kinase inhibition. Kinase activity assays with a panel of binding-deficient H-NOX mutants further reveal that while formation of the H-NOX-HK protein complex is required to stabilize the HK, H-NOX conformational changes upon NO binding are necessary for HK inhibition. Characterization of H-NOX proteins has led to an increased understanding of bacterial NO sensing. However, NO production in bacteria is less well-understood, and here the NOS protein from Synechococcus sp. PCC 7335 is characterized. Mammalian NOS proteins are comprised of a P450-like heme/oxidase domain responsible for catalysis, and a reductase domain responsible for electron transfer. While most bacterial NOS proteins discovered to date contain only the heme/oxidase domain, Synechococcus NOS contains both the oxidase and reductase domains, and additionally contains a predicted globin domain resembling bacterial flavohemoglobins. Spectroscopic and biochemical characterization of the globin indicated a possible role in redox communication in this novel class of bacterial NOS enzymes.

Download or read book Tonic and Acute Nitric Oxide Signaling Through Soluble Guanylate Cyclase written by Stephen Patrick Lewis Cary and published by . This book was released on 2005 with total page 450 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Biochemical Characterization of Oxygen Regulated Soluble Guanylate Cyclases written by Shirley Hsiao-Lin Huang and published by . This book was released on 2009 with total page 270 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Functional and Mechanistic Characterization of Bacterial H NOX Nitric Oxide Signaling Systems written by Lars Plate and published by . This book was released on 2013 with total page 141 pages. Available in PDF, EPUB and Kindle. Book excerpt: The gaseous free radical nitric oxide (NO) is firmly established as a unique signaling agent in nature. Eukaryotes employ this freely diffusible molecule in low transient concentrations as a cardiovascular signaling agent and neurotransmitter. Macrophages produce higher, cytotoxic concentrations of NO to serve as an integral piece of the host-defense against pathogens. NO signaling in vertebrates is well characterized and involves the Heme-Nitric oxide/Oxygen binding (H-NOX) domain of soluble guanylate cyclase (sGC) as a selective NO sensor. H-NOX domains are also present in many bacteria including a number of pathogens. Bacterial H-NOX proteins are often found in the same operon as signaling proteins such as histidine kinases, suggesting a role for H-NOX proteins as sensors in prokaryotic NO signaling pathways. H-NOX-dependent control of histidine kinase autophosphorylation has been demonstrated, but little was known about the biological role or output of H-NOX two-component signaling pathways in bacteria. Here, molecular details of a bacterial H-NOX signaling network in Shewanella oneidensis are presented. NO regulates biofilm formation by controlling the levels of the bacterial secondary messenger cyclic diguanosine monophosphate (cyclic-di-GMP) through an unusually complex multi-component signaling network. Homologous pathways exist in the pathogen Vibrio cholerae and in additional gammaproteobacteria. This work and other recent studies highlight a more general role for NO in restructuring bacterial communal behavior, influencing motility and biofilm formation, host-symbiont interactions, and quorum sensing. The connectivity of the signaling network in S. oneidensis was mapped by phosphotransfer profiling, which demonstrated signal integration from two histidine kinases and branching to three response regulators: HnoB, HnoC, and HnoD. Phosphodiesterase assays showed that a feed-forward loop between HnoB and HnoD response regulators with phosphodiesterase domains and phosphorylation-mediated activation intricately regulated c-di-GMP levels. In vivo phenotypic characterization established a direct link between NO signaling and increased biofilm formation. Cellular adhesion in biofilms may provide a general protection mechanism for bacteria against high concentrations of reactive and damaging NO. HnoC functions as transcription factor controlling expression of the signaling components in the network. Mechanistic studies of HnoC revealed an unprecedented regulation mechanism, involving phosphorylation-induced dissociation of the response regulator tetramer. The transcriptional feedback loop created by HnoC further regulates the dynamics of the H-NOX signaling network in response to NO stimuli. One of the phosphotransfer targets of the H-NOX-associated histidine kinase in the signaling network in S. oneidensis and V. cholerae contains a HD-GYP domain: a predicted but poorly characterized phosphodiesterase domain for cyclic-di-GMP hydrolysis. The HnoD HD-GYP domains contain degenerate residues that cause them to be catalytically inactive. To understand the functional consequence of the degeneracy and to gain general insights on the catalytic mechanism of HD-GYP domains, two catalytically active HD-GYP enzymes were characterized. The enzymes contained a binuclear iron center, and reconstitution experiments demonstrated that a heterovalent Fe(III)-Fe(II) cluster is likely required for catalysis. The absence of the metal-coordination site in HnoD eliminates any phosphodiesterase activity. The prevalence of orphan H-NOX/histidine kinase pairs highlights the necessity for improved methods to map connectivities in two-component signaling. The identity of cognate response regulator(s) is needed to define the biological function of the signaling system. Analogous to approaches in eukaryotic kinases, protein engineering was applied to histidine kinases to permit the use of unnatural ATP analogs as substrates, which are unreactive with other enzymes. Analog-sensitive alleles of two model histidine kinases were developed, and the phosphotransfer transfer reactions to their respective response regulators were optimized. Additionally, a panel of kinase inhibitors was screened for specificity against the analog-sensitive alleles. The analog-sensitive histidine kinases could become useful for in situ identification of phosphotransfer partners of orphan histidine kinases.

Download or read book Nitric Oxide Synthase Characterization and Functional Analysis written by and published by Academic Press. This book was released on 1996-09-18 with total page 375 pages. Available in PDF, EPUB and Kindle. Book excerpt: Although the function of nitric oxide in a regulatory capacity in the central and peripheral nervous system is widely recognized, the full scope of its actions and its interrelationships with other classes of regulatory molecules is just beginning to be comprehended. This volume contains a number of sophisticated and advanced methods essential for exploring the activity of nitric oxide in the brain. It will be a valuable tool for the established investigator and for those just entering the field. Comprehensive protocols included for detection of NO and related compounds by chemical, immunohistochemical, and in situ hybridization techniques Newly developed methods for the purification of neuronal and endothelial NO synthase, production of monoclonal antibodies to NO synthase, molecular cloning and expression of NO synthesis, and control of NO synthase gene expression Assessment of NO-mediated functions in neurons, central nervous system, cerebral circulation, synaptic transmission, and vascular tone Calcium imaging by confocal microscopy, evaluation of the effect of NO on iron metabolism, and detection of heme oxygenase-1 and -2 message level and distribution

Download or read book MOLECULAR MODEL OF SOLUBLE GUANYLYL CYCLASE INSIGHT INTO ALLOSTERY IN NITRIC OXIDE SIGNALING written by Bradley Fritz and published by . This book was released on 2011 with total page 386 pages. Available in PDF, EPUB and Kindle. Book excerpt: Soluble guanylyl cyclase (sGC), the nitric oxide (NO) receptor, is a 150 kDa heterodimeric multi-domain protein that contains heme in the beta subunit. Binding of NO to heme leads to rupture of the proximal histidine bond, increased catalytic conversion of GTP to cGMP at a distant guanylyl cyclase catalytic domain, and vasodilation through cGMP signaling. The structure of sGC has not been determined, and little is known about the mechanism by which NO binding to heme leads to increased catalysis. The small molecule YC-1 is known to stimulate sGC activity, but the exact YC-1 binding site and mechanism of action are unknown. Using truncated constructs of Manduca sexta (Ms) sGC lacking the catalytic domain, conformational changes upon YC-1 and NO-binding were characterized using analytical ultracentrifugation and small-angle X-ray scattering. Chemical cross-linking and high-resolution mass spectrometry was used to obtain distance restraints which, when combined with homology models, have provided the first model of sGC domain arrangement and revealed important information about domain-domain interactions. Truncated Ms sGC is highly elongated, contains a coiled-coil in a parallel arrangement, and contains a direct interface between the beta H-NOX (Heme Nitric oxide/Oxygen binding domain) and the coiled-coil and between the beta H-NOX and alpha PAS (Per-arnt-sim) domains. Experiments using analytical ultracentrifugation, fluorescence anisotropy and native mass spectrometry have revealed the YC-1 binding site to be located within the alpha PAS domain. Additionally, measurement of the kinetics of heme loss and the heme reduction potential were performed to investigate the instability of oxidized sGC heme.

Download or read book The Characterization of the Nitric Oxide cyclic GMP Pathway in the Auditory System written by James D. Fessenden and published by . This book was released on 1997 with total page 374 pages. Available in PDF, EPUB and Kindle. Book excerpt:

Download or read book Tuning Sensing and Signaling in the H NOX Family of Heme Proteins written by Michael B. Winter and published by . This book was released on 2011 with total page 238 pages. Available in PDF, EPUB and Kindle. Book excerpt: Nitric oxide (NO) signaling in mammals occurs through activation of the soluble isoform of guanylate cyclase (sGC), which results in diverse physiological processes such as blood vessel dilation and neurotransmission. sGC is a heterodimeric heme protein that has evolved to be a specific sensor for NO. Although sGC contains the same histidyl-ligated porphyrin as the globins, it has no measureable affinity for oxygen and does not oxidize in air. These unusual features facilitate selective and robust activation by NO in aerobic cellular contexts. Questions regarding the mechanism of ligand discrimination in sGC led to the identification of sGC heme domain homologues in many different organisms where these proteins function as NO and/or oxygen sensors for gas-mediated signaling pathways. The members of the protein family have been subsequently termed Heme Nitric oxide/OXygen binding (H-NOX) domains due to their divergent ligand-binding properties. Functional characterization of H-NOX domains from prokaryotes has provided important clues about the structural features that control ligand discrimination across the H-NOX family. However, additional fundamental questions remain about the influence of protein structure on heme chemistry. Extensive studies on the globins, as model heme proteins, have established the functional importance of heme protein topological features in modulating gas diffusion to and from the heme site. In globins, deep gas pockets around the porphyrin transiently capture and release oxygen, tuning oxygen affinity for transport, delivery, and storage in diverse physiological environments. Although mechanisms of gas binding in H-NOX proteins have been a subject of intense investigation, there is little knowledge regarding the functional role of protein structure in modulating gas diffusion. Using X-ray crystallography with xenon and kinetic measurements, a tunnel network that extends between the solvent and interior heme site was mapped in a prokaryotic H-NOX domain. Hindering gas diffusion through the tunnels has important consequences on diatomic gas affinity. This suggests that protein tunnels in H-NOX proteins may play functional roles in tuning gas-mediated signaling. Unlike isolated H-NOX domains, mammalian sGC is a structurally more complex, multi-domain protein. The ability of the sGC heme to resist oxidation in air is unique among histidyl-ligated heme proteins and essential for maximal NO-induced activation. Speculations have been made about how the sGC heme resists oxidation, but no study has systemically addressed the structural and electronic factors that contribute to this critical property. To probe the accessibility of oxygen to the heme site, we sought to substitute the native heme of sGC with an unnatural porphyrin displaying emission that is quenched by oxygen. Using an expression-based methodology, a phosphorescent Ru porphyrin was incorporated in sGC constructs of varying lengths. Emission quenching results suggest that oxygen diffusion to the full-length sGC heme site is significantly hindered compared to smaller sGC constructs. Limited oxygen accessibility, combined with heme electronic factors, appear to serve as important evolutionary solutions in sGC to protect the heme cofactor under aerobic conditions. Originally to probe sGC function, the strategies used to generate oxygen-sensing proteins have been applied to create a new family of H-NOX-based molecular sensors. Substitution of the native heme in bacterial H-NOX proteins with unnatural porphyrins has proven to be a promising strategy to develop stable protein agents for biological imaging applications. Heme proteins are highly tunable frameworks that coordinate porphyrins with high fidelity and specificity. These properties have been exploited to design heme protein-based sensors with tailored functionalities for enhancing porphyrin bioavailability. Initial work on making optical oxygen sensors has evolved into recent efforts to generate H-NOX-based MRI contrast agents with high relaxivities for future use in deep-tissue imaging. Thus, through substitution of the native heme group with unnatural porphyrins, new sensing properties have been built into the H-NOX family.

Download or read book Encyclopedia of Signaling Molecules written by Sangdun Choi and published by Springer. This book was released on 2017-12-15 with total page 6330 pages. Available in PDF, EPUB and Kindle. Book excerpt: The second edition of this encyclopedia presents over 400 biologically important signaling molecules and the content is built on the core concepts of their functions along with early findings written by some of the world’s foremost experts. The molecules are described by recognized leaders in each molecule. The interactions of these single molecules in signal transduction networks will also be explored. This encyclopedia marks a new era in overview of current cellular signaling molecules for the specialist and the interested non-specialist alike. Currently, there are more than 30,000 genes in human genome. However, not all the proteins encoded by these genes work equally in order to maintain homeostasis. Understanding the important signaling molecules as completely as possible will significantly improve our research-based teaching and scientific capabilities.

Download or read book Nitric Oxide Signaling Through Soluble Guanylate Cyclase written by and published by . This book was released on 2008 with total page 332 pages. Available in PDF, EPUB and Kindle. Book excerpt: Soluble guanylyl/guanylate cyclase (sGC), the primary receptor for nitric oxide (NO), is a heme containing heterodimeric enzyme involved in numerous physiological events in animals. The small molecule YC-1 stimulates sGC, but the mechanism behind and the location of binding are unknown. I have developed a prokaryotic expression system for insect (Manduca sexta) sGC. The recombinant holoenzyme, like its mammalian counterpart, is responsive to NO, CO and YC-1, displaying a 175-fold increase in activity on binding. Truncated constructs containing the N-terminal two-thirds of both subunits (msGC-NT) were designed to facilitate expression. With the highly pure material, we investigated NO and CO binding, reaction kinetics and regulation. Binding of NO to msGC-NT heme forms a six-coordinate intermediate followed by release of the proximal histidine to yield a five-coordinate nitrosyl complex. The conversion rate is insensitive to nucleotides, YC-1 and changes in NO concentration up to 3̃0 micromolar. In contrast, NO release from msGC-NT is biphasic in the absence of YC-1, while binding of YC-1 eliminates the fast phase but has little effect on the slower phase. CO binding to msGC-NT is also regulated by YC-1. The CO release rate is reduced by YC-1 while the on rate remains unchanged, which leads to an 5̃0-fold increase in binding affinity. YC-1 binding leads to a substantial geminate recombination of CO to msGC-NT upon photolysis. Our data are consistent with a model for allosteric activation in which (1) YC-1 binds away from the catalytic site and (2) sGC undergoes a conformational switch between two states of an open and a closed heme pocket. The final catalysis results from the integration of the influence of numerous allosteric effectors on the equilibrium between these two states.S-nitrosoglutathione (GSNO) existsin vivoand plays important roles in NO signaling. We have developed a model cell line, in which inducible NO synthase and human sGC genes were included. GSNO stimulation of sGC has been investigated using recombinant insect and human enzymes. GSNO can activate sGC as efficiently as gaseous NO, but apparently with a distinct mechanism. GSNO or endogenous NO couldS-nitrosylate sGC, which might regulate the enzyme function.

Download or read book Structural Genomics written by Yu Wai Chen and published by Humana. This book was released on 2013-11-09 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: The field of Structural Genomics has produced many technological advances that transform and accelerate structure solution and analysis. Structural Genomics: General Applications emphasizes the benefits to the wider structural research community. It also reflects the current trend in tackling the more ambitious challenges of studying macromolecular machineries and complexes. Divided into three convenient sections, topics include the cloning and production of proteins for structural studies, experimental methods, and computational methods and data analysis. Written in the successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible protocols, and notes on troubleshooting and avoiding known pitfalls. Authoritative and easily accessible, Structural Genomics: General Applications aims primarily to channel spin-off technologies to the average structural biologist in a small or medium-sized laboratory.

Download or read book The Physicochemical Aspects of Nitric Oxide in Chemistry and Biology written by Gertz I. Likhtenshtein and published by Nova Science Publishers. This book was released on 2018 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: Nitric oxide (NO) is a tiny molecule with tremendous impact in biology, biochemistry, physiology, biomedicine and clinical practice. Nitric oxide, as a signaling molecule, significantly contributes in the regulation of transcriptional, translational and post-translational processes, aging, inflammation, and major human pathologies. This book represents a view on the area from physical chemists with long and broad expertise in chemical biophysics, mechanisms of enzyme catalysis and physical methods for application in biology, in particular. This book forms the physicochemical basis for a deep understanding of the biochemical and physiological impact of nitric oxide. With numerous references to essential literature, this volume provides a tool to fundamental knowledge in all important aspects of the vast area. The book chapters are dedicated to basic physical and chemical properties of nitric oxide, NO donors, NO metallocomplexes, and methods of NO analysis. A general survey is made of data on nitric oxide synthase and soluble guanylate cyclase, an NO receptor, and reactive nitrogen species. Brief reviews of the nitric oxide involving genetics and aging, occurrence of pathology and treatment of cancer, arterial and bone diseases, central nervous systems and neurodegenerative diseases, and other miscellaneous diseases are all presented. This book is intended for scientists working in areas related to the impact of nitric oxide in biochemistry, molecular biology and biophysics, physiology and biomedicine. The book will also be useful for doctors and pharmacologists involved in the clinical practice. This text, as a whole or as separate chapters, can also be employed as subsidiary manuals for instructors, graduate and undergraduate students of university physics, bio-physics, chemistry, biochemistry, and biomedicine departments, and researchers of laboratories for pharmacological companies.

Download or read book Nitric Oxide written by Louis J. Ignarro and published by Academic Press. This book was released on 2000-09-13 with total page 1023 pages. Available in PDF, EPUB and Kindle. Book excerpt: Nitric oxide (NO) is a gas that transmits signals in an organism. Signal transmission by a gas that is produced by one cell and which penetrates through membranes and regulates the function of another cell represents an entirely new principle for signaling in biological systems. NO is a signal molecule of key importance for the cardiovascular system acting as a regulator of blood pressure and as a gatekeeper of blood flow to different organs. NO also exerts a series of other functions, such as acting a signal molecule in the nervous system and as a weapon against infections. NO is present in most living creatures and made by many different types of cells. NO research has led to new treatments for treating heart as well as lung diseases, shock, and impotence. Scientists are currently testing whether NO can be used to stop the growth of cancerous tumors, since the gas can induce programmed cell death, apoptosis. This book is the first comprehensive text on nitric oxide to cover all aspects--basic biology, chemistry, pathobiology, effects on various disease states, and therapeutic implications. Edited by Nobel Laureate Louis J. Ignarro, editor of the Academic Press journal, Nitric Oxide Authored by world experts on nitric oxide Includes an overview of basic principles of biology and chemical biology Covers principles of pathobiology, including the nervous system, cardiovascular function, pulmonary function, and immune defense

Download or read book Essentials of Medical Biochemistry written by Chung Eun Ha and published by Academic Press. This book was released on 2011-01-28 with total page 600 pages. Available in PDF, EPUB and Kindle. Book excerpt: Expert biochemist N.V. Bhagavan’s new work condenses his successful Medical Biochemistry texts along with numerous case studies, to act as an extensive review and reference guide for both students and experts alike. The research-driven content includes four-color illustrations throughout to develop an understanding of the events and processes that are occurring at both the molecular and macrolecular levels of physiologic regulation, clinical effects, and interactions. Using thorough introductions, end of chapter reviews, fact-filled tables, and related multiple-choice questions, Bhagavan provides the reader with the most condensed yet detailed biochemistry overview available. More than a quick survey, this comprehensive text includes USMLE sample exams from Bhagavan himself, a previous coauthor. Clinical focus emphasizing relevant physiologic and pathophysiologic biochemical concepts Interactive multiple-choice questions to prep for USMLE exams Clinical case studies for understanding basic science, diagnosis, and treatment of human diseases Instructional overview figures, flowcharts, and tables to enhance understanding