共查询到20条相似文献,搜索用时 750 毫秒
1.
J Al-Hijji Ellika Andolf Ricardo Laurini Satish Batra 《Reproductive biology and endocrinology : RB&E》2003,1(1):51
To investigate the possible role of nitric oxide (NO) produced locally or intramurally in the quiescence of the pregnant myometrium,
nitric oxide synthase (NOS) activity was measured in samples from first trimester (villous, and non villous-trophoblast),
term placenta and pregnant myometrium. Trophoblast tissue was obtained from psychosocial termination of pregnancy (9 – 12
weeks' gestation) whereas placenta and myometrium, from the same patient, at deliveries by Caesarean section. NOS activity
was measured in both cytosolic and particulate fractions by the formation of 14C-citrulline from 14C-arginine. Western immunoblotting was used to identify the endothelial NOS (eNOS) and neuronal (nNOS) isoforms. The activity
of NOS in particulate fractions from all preparations was considerably higher than the cytosolic fractions. Activity in all
fractions except the myometrium was highly Ca-dependent. More than 50% of particulate NOS from the myometrium was Ca-independent.
NOS activity was highest in the villous trophoblast and there was a significant difference between the villous and non-villous
trophoblast. In placenta and myometrium, NOS was 2–4 fold and 20–28-fold lower than the villous trophoblast, respectively.
Western blot analysis showed clearly eNOS in the particulate fraction and a weak eNOS band in the cytosolic fractions, whereas
nNOS was not detectable in any of the fractions. In view of the marginal activity of NOS in the myometrium, NO produced by
the trophoblast and placenta could play a significant role in maintaining uterine quiescence by paracrine effect. 相似文献
2.
Flinspach ML Li H Jamal J Yang W Huang H Hah JM Gómez-Vidal JA Litzinger EA Silverman RB Poulos TL 《Nature structural & molecular biology》2004,11(1):54-59
Three nitric oxide synthase (NOS) isoforms, eNOS, nNOS and iNOS, generate nitric oxide (NO) crucial to the cardiovascular, nervous and host defense systems, respectively. Development of isoform-selective NOS inhibitors is of considerable therapeutic importance. Crystal structures of nNOS-selective dipeptide inhibitors in complex with both nNOS and eNOS were solved and the inhibitors were found to adopt a curled conformation in nNOS but an extended conformation in eNOS. We hypothesized that a single-residue difference in the active site, Asp597 (nNOS) versus Asn368 (eNOS), is responsible for the favored binding in nNOS. In the D597N nNOS mutant crystal structure, a bound inhibitor switches to the extended conformation and its inhibition of nNOS decreases >200-fold. Therefore, a single-residue difference is responsible for more than two orders of magnitude selectivity in inhibition of nNOS over eNOS by L-N(omega)-nitroarginine-containing dipeptide inhibitors. 相似文献
3.
Neuronal nitric-oxide synthase (nNOS or NOS I) and endothelial NOS (eNOS or NOS III) differ widely in their reductase and nitric oxide (NO) synthesis activities, electron transfer rates, and propensities to form a heme-NO complex during catalysis. We generated chimeras by swapping eNOS and nNOS oxygenase domains to understand the basis for these differences and to identify structural elements that determine their catalytic behaviors. Swapping oxygenase domains did not alter domain-specific catalytic functions (cytochrome c reduction or H(2)O(2)-supported N(omega)-hydroxy-l-arginine oxidation) but markedly affected steady-state NO synthesis and NADPH oxidation compared with native eNOS and nNOS. Stopped-flow analysis showed that reductase domains either maintained (nNOS) or slightly exceeded (eNOS) their native rates of heme reduction in each chimera. Heme reduction rates were found to correlate with the initial rates of NADPH oxidation and heme-NO complex formation, with the percentage of heme-NO complex attained during the steady state, and with NO synthesis activity. Oxygenase domain identity influenced these parameters to a lesser degree. We conclude: 1) Heme reduction rates in nNOS and eNOS are controlled primarily by their reductase domains and are almost independent of oxygenase domain identity. 2) Heme reduction rate is the dominant parameter controlling the kinetics and extent of heme-NO complex formation in both eNOS and nNOS, and thus it determines to what degree heme-NO complex formation influences their steady-state NO synthesis, whereas oxygenase domains provide minor but important influences. 3) General principles that relate heme reduction rate, heme-NO complex formation, and NO synthesis are not specific for nNOS but apply to eNOS as well. 相似文献
4.
In a continuing effort to unravel the structural basis for isoform-selective inhibition of nitric oxide synthase (NOS) by various inhibitors, we have determined the crystal structures of the nNOS and eNOS heme domain bound with two D-nitroarginine-containing dipeptide inhibitors, D-Lys-D-Arg(NO)2-NH(2) and D-Phe-D-Arg(NO)2-NH(2). These two dipeptide inhibitors exhibit similar binding modes in the two constitutive NOS isozymes, which is consistent with the similar binding affinities for the two isoforms as determined by K(i) measurements. The D-nitroarginine-containing dipeptide inhibitors are not distinguished by the amino acid difference between nNOS and eNOS (Asp 597 and Asn 368, respectively) which is key in controlling isoform selection for nNOS over eNOS observed for the L-nitroarginine-containing dipeptide inhibitors reported previously [Flinspach, M., et al. (2004) Nat. Struct. Mol. Biol. 11, 54-59]. The lack of a free alpha-amino group on the D-nitroarginine moiety makes the dipeptide inhibitor steer away from the amino acid binding pocket near the active site. This allows the inhibitor to extend into the solvent-accessible channel farther away from the active site, which enables the inhibitors to explore new isoform-specific enzyme-inhibitor interactions. This might be the structural basis for why these D-nitroarginine-containing inhibitors are selective for nNOS (or eNOS) over iNOS. 相似文献
5.
Isao Yokoi Yukiko Namba Hideaki Kabuto Kazunori Inada Motoyuki Iida Akitane Mori Norio Ogawa 《Neurochemical research》1996,21(10):1187-1192
Since nitric oxide (NO) is synthesized by nitric oxide synthase (NOS) froml-arginine (Arg) which has an amidino group in its molecule, we, examined the effect of 29 kinds of Arg analogues on neuronal
NOS (nNOS) activity in the rat brain. None of the Arg analogues acted as a substrate for nNOS. Diamidinocystamine, hirudonine,
and guanethidine inhibited nNOS activity to 67.3%, 64.2% and 74.1%, respectively, but their inhibitory efficiency was lower
than NG-monomethyl-l-arginine (to 36.5%) which is a well known NOS inhibitor. Dimethylguanidine and N-benzoylguanidine also significantly inhibited
nNOS activity to 88.0% and 90.7%, respectively. Whereas almost all of the NOS inhibitors previously reported were synthesizdd
by substituting the amidino nitrogen of Arg, none of these new inhibitors were substituted at this position. Furthermore,
hirudonine, which is a naturally occurring compound, was thought to act as an agonist at polyamine binding site of the N-methyl-d-aspartate type of glutamate receptor complex. It is also interesting that guanethidine, an antihypertensive agent, inhibit
nNOS activity. These new drugs are useful for the investigation not only of the chemical nature of nNOS but also of the physiologic
function of NO. 相似文献
6.
Qing Jing Huiying Li Jianguo Fang Linda J. Roman Pavel Martásek Thomas L. Poulos Richard B. Silverman 《Bioorganic & medicinal chemistry》2013,21(17):5323-5331
In certain neurodegenerative diseases damaging levels of nitric oxide (NO) are produced by neuronal nitric oxide synthase (nNOS). It, therefore, is important to develop inhibitors selective for nNOS that do not interfere with other NOS isoforms, especially endothelial NOS (eNOS), which is critical for proper functioning of the cardiovascular system. While we have been successful in developing potent and isoform-selective inhibitors, such as lead compounds 1 and 2, the ease of synthesis and bioavailability have been problematic. Here we describe a new series of compounds including crystal structures of NOS-inhibitor complexes that integrate the advantages of easy synthesis and good biological properties compared to the lead compounds. These results provide the basis for additional structure–activity relationship (SAR) studies to guide further improvement of isozyme selective inhibitors. 相似文献
7.
Diaz-Ruiz A Vergara P Perez-Severiano F Segovia J Guizar-Sahagún G Ibarra A Ríos C 《Neurochemical research》2005,30(2):245-251
Nitric oxide (NO) plays a role in the pathophysiology of spinal cord injury (SCI). NO is produced by three types of nitric oxide synthase (NOS) enzymes: The constitutive Ca2+/calmodulin-dependent neuronal NOS (nNOS) and endothelial NOS (eNOS) isoforms, and the inducible calcium-independent isoform (iNOS). During the early stages of SCI, nNOS and eNOS produce significant amounts of NO, therefore, the regulation of their activity and expression may participate in the damage after SCI. In the present study, we used Cyclosporin-A (CsA) to further substantiate the role of Ca-dependent NOS in neural responses associated to SCI. Female Wistar rats were subjected to SCI by contusion, and killed 4 h after lesion. Results showed an increase in the activity of constitutive NOS (cNOS) after lesion, inhibited by CsA (2.5 mg/kg i.p.). Western blot assays showed an increased expression of both nNOS and eNOS after trauma, also antagonized by CsA administration. 相似文献
8.
Li H Shimizu H Flinspach M Jamal J Yang W Xian M Cai T Wen EZ Jia Q Wang PG Poulos TL 《Biochemistry》2002,41(47):13868-13875
A series of N-alkyl-N'-hydroxyguanidine compounds have recently been characterized as non-amino acid substrates for all three nitric oxide synthase (NOS) isoforms which mimic NO formation from N(omega)-hydroxy-L-arginine. Crystal structures of the nNOS heme domain complexed with either N-isopropyl-N'-hydroxyguanidine or N-butyl-N'-hydroxyguanidine reveal two different binding modes in the substrate binding pocket. The binding mode of the latter is consistent with that observed for the substrate N(omega)-hydroxy-L-arginine bound in the nNOS active site. However, the former binds to nNOS in an unexpected fashion, thus providing new insights into the mechanism on how the hydroxyguanidine moiety leads to NO formation. Structural features of substrate binding support the view that the OH-substituted guanidine nitrogen, instead of the hydroxyl oxygen, is the source of hydrogen supplied to the active ferric-superoxy species for the second step of the NOS catalytic reaction. 相似文献
9.
Holmqvist B Olsson CF Svensson ML Svanborg C Forsell J Alm P 《Journal of molecular histology》2005,36(8-9):499-516
We determined the cellular mRNA expression of all intrarenal nitric oxide (NO)-producing NO synthase (NOS) isoforms, endothelial
NOS (eNOS) and neuronal NOS (nNOS) and inducible NOS (iNOS) in kidneys from wild-type mice (WT) and immune deficient Toll-like
receptor 4 (TLR4) mutant mice, during normal physiological conditions and during a short-term (6–16 h) endotoxic condition
caused by systemically administered lipopolysaccaride (LPS). Investigations were performed by means of in situ hybridization and polymerase chain reaction amplification techniques. In WT, LPS altered the expression rate of all intrarenal
NOS isoforms in a differentiated but NOS-isoform coupled expression pattern, with iNOS induction, and up- and down-regulation
of the otherwise constitutively expressed NOS isoforms, e.g. eNOS and nNOS and an iNOS isotype. In TLR4 mutants, LPS caused
none or a lowered iNOS induction, but altered the expression rate of the constitutive NOS isoforms. It is concluded that the
intrarenal spatial relation of individual NOS-isoforms and their alteration in expression provide the basis for versatile
NO-mediated renal actions that may include local interactions between NOS isoforms and their individual NO-target sites, and
that the NOS-isoform dependent events are regulated by TLR4 during endotoxic processes. These regulatory mechanisms are likely
to participate in different pathophysiological conditions affecting NO-mediated renal functions. 相似文献
10.
Le Bourdonnec B Leister LK Ajello CA Cassel JA Seida PR O'Hare H Gu M Chu GH Tuthill PA DeHaven RN Dolle RE 《Bioorganic & medicinal chemistry letters》2008,18(1):336-343
Nitric oxide (NO), a mediator of various physiological and pathophysiological processes, is synthesized by three isozymes of nitric oxide synthase (NOS). Potential candidate clinical drugs should be devoid of inhibitory activity against endothelial NOS (eNOS), since eNOS plays an important role in maintaining normal blood pressure and flow. A new series of aminopiperidines as potent inhibitors of iNOS were identified from a HTS lead. From this study, we identified compound 33 as a potent iNOS inhibitor, with >25-fold selectivity over eNOS and 16-fold selectivity over nNOS. 相似文献
11.
Hojná S Kuneš J Zicha J 《Physiological research / Academia Scientiarum Bohemoslovaca》2010,59(6):997-1009
Both brain and peripheral nitric oxide (NO) play a role in the control of blood pressure and circulatory homeostasis. Central NO production seems to counteract angiotensin II-induced enhancement of sympathetic tone. The aim of our study was to evaluate NO synthase (NOS) activity and protein expression of its three isoforms--neuronal (nNOS), endothelial NOS (eNOS) and inducible (iNOS)--in two brain regions involved in blood pressure control (diencephalon and brainstem) as well as in the kidney of young adult rats with either genetic (12-week-old SHR) or salt-induced hypertension (8-week-old Dahl rats). We have demonstrated reduced nNOS and iNOS expression in brainstem of both hypertensive models. In SHR this abnormality was accompanied by attenuated NOS activity and was corrected by chronic captopril treatment which prevented the development of genetic hypertension. In salt hypertensive Dahl rats nNOS and iNOS expression was also decreased in the diencephalon where neural structures important for salt hypertension development are located. As far as peripheral NOS activity and expression is concerned, renal eNOS expression was considerably reduced in both genetic and salt-induced hypertension. In conclusions, we disclosed similar changes of NO system in the brainstem (but not in the diencephalon) of rats with genetic and salt-induced hypertension. Decreased nNOS expression was associated with increased blood pressure due to enhanced sympathetic tone. 相似文献
12.
Saluja R Jyoti A Chatterjee M Habib S Verma A Mitra K Barthwal MK Bajpai VK Dikshit M 《Biochimica et biophysica acta》2011,1813(10):1700-1707
13.
An-Soo Jang Inseon-S Choi Jong-Un Lee Sung-Woo Park June-Hyuk Lee Choon-Sik Park 《Respiratory research》2004,5(1):5
BackgroundThe functional role of nitric oxide (NO) and various nitric oxide synthase (NOS) isoforms in asthma remains unclear.ObjectiveThis study investigated the effects of ozone and ovalbumin (OVA) exposure on NOS isoforms.MethodsThe expression of inducible NOS (iNOS), neuronal NOS (nNOS), and endothelial NOS (eNOS) in lung tissue was measured. Enhanced pause (Penh) was measured as a marker of airway obstruction. Nitrate and nitrite in bronchoalveolar lavage (BAL) fluid were measured using a modified Griess reaction.ResultsThe nitrate concentration in BAL fluid from the OVA-sensitized/ozone-exposed/OVA-challenged group was greater than that of the OVA-sensitized/saline-challenged group. Methacholine-induced Penh was increased in the OVA-sensitized/ozone-exposed/OVA-challenged group, with a shift in the dose-response curve to the left, compared with the OVA-sensitized/saline-challenged group. The levels of nNOS and eNOS were increased significantly in the OVA-sensitized/ozone-exposed/OVA-challenged group and the iNOS levels were reduced compared with the OVA-sensitized/saline-challenged group.ConclusionIn mice, ozone is associated with increases in lung eNOS and nNOS, and decreases in iNOS. None of these enzymes are further affected by allergens, suggesting that the NOS isoforms play different roles in airway inflammation after ozone exposure. 相似文献
14.
Vaughan DJ Brogan TV Kerr ME Deem S Luchtel DL Swenson ER 《American journal of physiology. Lung cellular and molecular physiology》2003,284(5):L834-L843
We investigated the source(s) for exhaled nitric oxide (NO) in isolated, perfused rabbits lungs by using isozyme-specific nitric oxide synthase (NOS) inhibitors and antibodies. Each inhibitor was studied under normoxia and hypoxia. Only nitro-L-arginine methyl ester (L-NAME, a nonselective NOS inhibitor) reduced exhaled NO and increased hypoxic pulmonary vasoconstriction (HPV), in contrast to 1400W, an inhibitor of inducible NOS (iNOS), and 7-nitroindazole, an inhibitor of neuronal NOS (nNOS). Acetylcholine-mediated stimulation of vascular endothelial NOS (eNOS) increased exhaled NO and could only be inhibited by L-NAME. Selective inhibition of airway and alveolar epithelial NO production by nebulized L-NAME decreased exhaled NO and increased hypoxic pulmonary artery pressure. Immunohistochemistry demonstrated extensive staining for eNOS in the epithelia, vasculature, and lymphatic tissue. There was no staining for iNOS but moderate staining for nNOS in the ciliated cells of the epithelia, lymphoid tissue, and cartilage cells. Our findings show virtually all exhaled NO in the rabbit lung is produced by eNOS, which is present throughout the airways, alveoli, and vessels. Both vascular and epithelial-derived NO modulate HPV. 相似文献
15.
Ellenberger EA Lucas HL Mueller JL Barrington PL Chung E Ohgami Y Quock RM 《Life sciences》2004,76(5):499-507
This study was conducted to demonstrate the involvement of nitric oxide synthase (NOS) in the early-phase isoflurane-induced hypotension and to ascertain whether this NOS is neuronal NOS (nNOS) or endothelial NOS (eNOS). Mean arterial pressures (MAPs) were directly measured from the femoral arteries of urethane-anesthetized rats. Isoflurane-induced changes in MAP were monitored in rats following pretreatment with vehicle or one of the following NOS inhibitors: L-NG-monomethyl-L-arginine (L-NMMA), which is non-selective; L-NG-nitro arginine (L-NOARG), which is more selective for nNOS and eNOS; and 7-nitroindazole (7-NI), which is selective for nNOS. Exposure to 2% isoflurane in oxygen produced a triphasic reduction in MAP, including an early phase in which mean arterial pressure (MAP) fell by 25-30% during the initial 2½ min. This early hypotensive response, but not subsequent phases, was abolished by i.v. pretreatment with either L-NMMA or L-NOARG. The early-phase hypotension was also significantly attenuated by i.p. pretreatment with 7-NI; however, the blockade was not as complete as with L-NMMA or L-NOARG. Cerebella and aorta were removed from vehicle- and 7-NI pretreated rats and assayed for NOS activity by determining the conversion of [14C]L-arginine to [14C]L-citrulline. The 7-NI pretreatment significantly reduced NOS activity in the cerebellum but not the aorta. These findings indicate that the early-phase isoflurane-induced hypotension may involve nNOS as well as eNOS. The nNOS may participate in regulation of isoflurane-induced neuronal release of endogenous opioid peptide, which produces a vasodilation that is dependent on NO derived from an action of eNOS. 相似文献
16.
17.
Fagan KA McMurtry I Rodman DM 《Physiological research / Academia Scientiarum Bohemoslovaca》2000,49(5):539-548
Nitric oxide (NO) is implicated in a wide variety of biological roles. NO is generated from three nitric oxide synthase (NOS) isoforms: neuronal (nNOS), inducible (iNOS), and endothelial (eNOS) all of which are found in the lung. While there are no isoform-specific inhibitors of NOS, the recent development and characterization of mice deficient in each of the NOS isoforms has allowed for more comprehensive study of the importance of NO in the lung circulation. Studies in the mouse have identified the role of NO from eNOS in modulating pulmonary vascular tone and in attenuating the development of chronic hypoxic pulmonary hypertension. 相似文献
18.
Huiying Li Fengtian Xue James M. Kraus Haitao Ji Kristin Jansen Labby Jan Mataka Silvia L. Delker Pavel Martásek Linda J. Roman Thomas L. Poulos Richard B. Silverman 《Bioorganic & medicinal chemistry》2013,21(5):1333-1343
Inhibitors of neuronal nitric oxide synthase have been proposed as therapeutics for the treatment of different types of neurological disorders. On the basis of a cis-3,4-pyrrolidine scaffold, a series of trans-cyclopropyl- and methyl-containing nNOS inhibitors have been synthesized. The insertion of a rigid electron-withdrawing cyclopropyl ring decreases the basicity of the adjacent amino group, which resulted in decreased inhibitory activity of these inhibitors compared to the parent compound. Nonetheless, three of them exhibited double-digit nanomolar inhibition with high nNOS selectivity on the basis of in vitro enzyme assays. Crystal structures of nNOS and eNOS with these inhibitors bound provide a basis for detailed structure–activity relationship (SAR) studies. The conclusions from these studies will be used as a guide in the future development of selective NOS inhibitors. 相似文献
19.
Schmidt K Andrew P Schrammel A Groschner K Schmitz V Kojda G Mayer B 《American journal of physiology. Heart and circulatory physiology》2001,281(5):H2053-H2061
The neuronal and endothelial isoforms of nitric oxide (NO) synthase (nNOS and eNOS, respectively) both catalyze the production of NO but are regulated differently. Stably transfected HEK 293 cell lines containing nNOS, eNOS, and a soluble mutant of eNOS were therefore established to compare their activity in a common cellular environment. NOS activity was determined by measuring L-[3H]citrulline production in homogenates and intact cells, the conversion of oxyhemoglobin to methemoglobin, and the production of cGMP. The results indicate that nNOS is more active than eNOS, both in unstimulated as well as calcium-stimulated cells. Under basal conditions, the soluble mutant of eNOS appeared to be slightly more active than wild-type eNOS in terms of NO and cGMP formation, suggesting that membrane association may be crucial for inhibition of basal NO release but is not required for stimulation by Ca2+-mobilizing agents. The maximal activity of soluble guanylate cyclase was significantly reduced by transfection with wild-type eNOS due to downregulation of mRNA expression. These results demonstrate that nNOS and eNOS behave differently even in an identical cellular environment. 相似文献