Showing papers on "Nitrosylation published in 1997"

Caspase-mediated apoptosis in neuronal excitotoxicity triggered by nitric oxide.

Abstract: Excitotoxicity and excess generation of nitric oxide (NO) are believed to be fundamental mechanisms in many acute and chronic neurodegenerative disorders. Disturbance of Ca2+ homeostasis and protein nitration/nitrosylation are key features in such conditions. Recently, a family of proteases collectively known as caspases has been implicated as common executor of a variety of death signals. In addition, overactivation of poly-(ADP-ribose) polymerase (PARP) has been observed in neuronal excitotoxicity. We therefore designed this study to investigate whether triggering of caspase activity and/or activation of PARP played a role in cerebellar granule cell (CGC) apoptosis elicited by peroxynitrite (ONOO−) or NO donors. CGC from wild-type or PARP −/− mice were exposed to various nitric oxide donors. Caspase activation and its implications for membrane alterations, Ca2+ homeostasis, intracellular proteolysis, chromatin degradation, and cell death were investigated. CGC exposed to NO donors undergo apoptosis, which is mediated by excess synaptic release of excitotoxic mediators. This excitotoxic mechanism differs from direct NO toxicity in some other neuronal populations and does not involve PARP activation. Inhibition of caspases with different peptide substrates prevented cell death and the related features, including intracellular proteolysis, chromatin breakdown, and translocation of phosphatidylserine to the outer surface of the cell membrane. Increased Ca2+ influx following N-methyl-D-aspartate (NMDA) receptor (NMDA-R) activation was not inhibited by caspase inhibitors. In CGC, NO donors elicit apoptosis by a mechanism involving excitotoxic mediators, Ca2+ overload, and subsequent activation of caspases.

Nitrosylated bovine serum albumin derivatives as pharmacologically active nitric oxide congeners

Abstract: Although nitrosothiols have been suggested to act as regulators of cell (patho)physiology, little is known about the pharmacology of nitrosylated proteins as nitric oxide (NO ⋅ ) congeners. We describe the molecular consequences of nitrosylating bovine serum albumin (BSA) at multiple specific sites and demonstrate that the product S -nitrosoproteins exert NO ⋅ -like activity. The content of nucleophilic nitrosylation sites ( i.e., free sulfhydryl groups) in native BSA was increased by either reduction with dithiothreitol or thiolation with N-acetylhomocysteine. Fourteen moles of nitrogen monoxide (NO)/mol BSA equivalent were then selectively positioned on either the endogenous sulfhydryl groups of reduced BSA or the homocysteine moieties of thiolated BSA, respectively. Each resulting S -nitrosoprotein adduct was an oligomeric mixture across the >2000 kDa to ≈66 kDa molecular mass range. The BSA-derived S -nitrosoproteins were immunoreactive with antibodies against native BSA but evidenced compromised long-chain fatty acid binding. Both types of BSA-derived S -nitrosoproteins suppressed human coronary artery smooth muscle cell proliferation to a similar degree (IC 50 ≈70 μM NO ⋅ equivalents) and were significantly more effective antiproliferative agents than a standard NO ⋅ donor, DETA NONOate. Antiproliferative bioactivity reflected the NO functionalities carried by each protein, but was independent of molecular mass of the nitrosylated BSA adducts. These data exemplify the rational design and characterization of protein-based S -nitrosothiols as NO ⋅ congeners and suggest that such agents could have therapeutic potential as NO delivery systems.

Regulation of cellular thiol redox status by nitric oxide.

Abstract: Nitric oxide (NO) is an important biological molecule that participates in a wide range of responses, including vasodilation, platelet regulation, neurotransmission, and cytoxicity (1). NO’s most widely known and best understood mechanism of action is through its interaction with the heme group in proteins such as soluble guanylate cyclase (1). In addition to heme iron, NO has also been shown to react with other functional groups in proteins. However, it is becoming clear that many reactions previously attributed to NO are now known to be mediated through NO-derived species (NOx) and not directly through NO itself (as discussedbelow). The thiol group of cysteine residues represents one potential target of NOx and may be of particular biological relevance since cysteines are important for maintaining the native conformation of proteins, are critical to the activity of many enzymes, and are the most reactive amino acid residues at physiological pH. In fact, modification of protein thiols by NOx alters the function and activity of various proteins including enzymes, signaling proteins, ion channels, receptors, transcription factors, and antioxidants (2–11). These alterations in protein function, through modification of thiols, may be critical for initiating signaling events or may be detrimental to the cell by disrupting essential protein function. This article aims to review the interactions of NOx with protein and nonprotein thiols, including mechanisms of thiol modification, reversibility of NOx-mediated protein thiol modification, and cellular defense mechanisms involved in regulating NOx-mediated modification of protein thiols.

60 – NO-Related Species: Neuroprotection versus Neurodestruction

Abstract: This chapter outlines the important roles of different NO-related species: nitric oxide (NO·), redox-related form of the NO group with one less electron (NO + ), and redox-related form of the NO group with one additional electron (NO – ). Recent evidence suggests that all three of these redox-related forms are pharmacologically important and also represent physiological forms of the NO group participating in distinctive chemical reactions. The actions of the NO group are correlated with its redox state. The NO· appears to be associated with neuronal injury caused by excessive activation of the NMDA receptor. In this scenario, peroxynitrite is formed from the reaction of nitric oxide (NO·) with superoxide anion (O 2 · − ). Drugs containing the NO group in alternative redox states may prevent NMDA receptor-mediated neurotoxicity. In the case of NO + equivalents, this mechanism appears to involve S -nitrosylation or oxidation of critical thiols to disulfide bonds in the NMDA receptor's redox modulatory site(s) to downregulate channel activity. Recent data also suggest that NO − , probably in the singlet state, can react with critical sulfhydryl group(s) of the NMDA receptor to downregulate its activity.

Caspase-Mediated Apoptosis in NeuronalExcitotoxicity Triggered byNitricOxide

Abstract: Background: Excitotoxicity andexcess generation of nitric oxide (NO)arebelieved tobefundamental mechanisms inmanyacuteandchronic neurodegenerative disorders. Disturbance ofCa2" homeostasis andprotein nitration/nitrosylation arekeyfeatures insuchconditions. Recently, afamily ofproteases collectively known ascaspases hasbeenimplicated ascommonexecutor of avariety ofdeath signals. Inaddition, overactivation of poly-(ADP-ribose) polymerase (PARP)hasbeenobserved inneuronal excitotoxicity. Wetherefore designed this study toinvestigate whether triggering ofcaspase activity and/or activation ofPARPplayed arole incerebellar granule cell (CGC)apoptosis elicited byperoxynitrite (ONOO-)orNO donors. Materials andMethods: CGCfromwild-type orPARP -/-micewereexposed tovarious nitric oxide donors. Caspase activation anditsimplications formembrane alterations, Ca2+homeostasis, intracellular proteolysis, chromatin degradation, andcell death wereinvestigated. Results: CGCexposed toNOdonors undergo apoptosis, whichismediated byexcess synaptic release ofexcitotoxic mediators. This excitotoxic mechanism differs from direct NO toxicity insomeother neuronal populations anddoesnotinvolve PARPactivation. Inhibition of caspases withdifferent peptide substrates prevented cell deathandtherelated features, including intracellular proteolysis, chromatin breakdown, andtranslocation of phosphatidylserine totheouter surface ofthecell membrane. Increased Ca2" influx following N-methyl-D-aspartate (NMDA)receptor (NMDA-R)activation wasnot inhibited bycaspase inhibitors. Conclusions: InCGC,NO donors elicit apoptosis bya mechanism involving excitotoxic mediators, Ca2+overload, andsubsequent activation ofcaspases.