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S Malek

Publications and source records attributed to S Malek.

16 recordsLinked to original sources
PubMed2000

Preparation and crystallization of dynamic NF-kappa B.Ikappa B complexes.

The formation of single, well-diffracting crystals is a requirement for any molecular structure determination by x-ray crystallography. Crystallization of biological macromolecules can represent a significant obstacle when the subject exhibits internal flexibility or indiscriminate self-association. In such cases, the removal of inherently flexible regions and the addition of stabilizing ligands can improve the probability of crystal formation and ordered growth. We have applied these principles in order to form crystals of the Rel homology region of transcription factor NF-kappaB in complex with its inhibitors IkappaBalpha and IkappaBbeta. None of these molecules crystallizes in the absence of a binding partner. Recombinant overexpression of truncated IkappaBalpha required selection of the correct start site. NF-kappaB.IkappaBalpha complex crystals formed under relatively stringent conditions. NF-kappaB. IkappaBbeta complex crystals were formed by analogy to NF-kappaB. IkappaBalpha, although some modifications in purification and complex formation were necessary due to differences between the inhibitors.

Animals↗
PubMed2000

Mechanism of kappa B DNA binding by Rel/NF-kappa B dimers.

The DNA binding of three different NF-kappaB dimers, the p50 and p65 homodimers and the p50/p65 heterodimer, has been examined using a combination of gel mobility shift and fluorescence anisotropy assays. The NF-kappaB p50/p65 heterodimer is shown here to bind the kappaB DNA target site of the immunoglobulin kappa enhancer (Ig-kappaB) with an affinity of approximately 10 nm. The p50 and p65 homodimers bind to the same site with roughly 5- and 15-fold lower affinity, respectively. The nature of the binding isotherms indicates a cooperative mode of binding for all three dimers to the DNA targets. We have further characterized the role of pH, salt, and temperature on the formation of the p50/p65 heterodimer-Ig-kappaB complex. The heterodimer binds to the Ig-kappaB DNA target in a pH-dependent manner, with the highest affinity between pH 7.0 and 7.5. A strong salt-dependent interaction between Ig-kappaB and the p50/p65 heterodimer is observed, with optimum binding occurring at monovalent salt concentrations below 75 mm, with binding becoming virtually nonspecific at a salt concentration of 200 mm. Binding of the heterodimer to DNA was unchanged across a temperature range between 4 degrees C and 42 degrees C. The sensitivity to ionic environment and insensitivity to temperature indicate that NF-kappaB p50/p65 heterodimers form complexes with specific DNA in an entropically driven manner.

Amino Acid Substitution↗
PubMed2000

[Cardiogenic shock in the acute phase of myocardial infarction].

Cardiogenic shock (CC) is the most common cause of death in acute myocardial infarction (MI) and it has been often associated with fatal evolution. The aim of this study is to emphasize the positive impact of myocardial revascularisation especially percutaneous coronary angioplasty (PTCA) on short and mid term survival of 35 patients hospitalised for acute MI with CC. All the 11 patients who were on conventional therapy died, whereas the hospital mortality rate of the 24 patients who underwent primary PTCA was 41%. PTCA in CC complicating MI is highly effective and reduces the mortality rate in the literature from 80% to 40%. For this reason PTCA should be performed in each case of MI with CC seen in the first 6 hours.

Aged↗
PubMed1998

Immunostimulatory DNA sequences inhibit IL-5, eosinophilic inflammation, and airway hyperresponsiveness in mice.

We have used a mouse model of allergen-induced airway hyperresponsiveness to demonstrate that immunostimulatory DNA sequences (ISS) containing a CpG DNA motif significantly inhibit airway eosinophilia and reduce responsiveness to inhaled methacholine. ISS not only inhibited eosinophilia of the airway (by 93%) and lung parenchyma (91%), but also significantly inhibited blood eosinophilia (86%), suggesting that ISS was exerting a significant effect on the bone marrow production of eosinophils. The inhibition of the bone marrow production of eosinophils by 58% was associated with a significant inhibition of T cell-derived cytokine generation (IL-5, granulocyte-macrophage CSF, and IL-3). ISS exerted this inhibitory effect on T cell cytokine production indirectly by stimulating monocytes/macrophages and NK cells to generate IL-12 and IFNs. The onset of the ISS effect on reducing the number of tissue eosinophils was both immediate (within 1 day of administration) and sustained (lasted 6 days), and was not due to ISS directly inducing eosinophil apoptosis. ISS was effective in inhibiting eosinophilic airway inflammation when administered either systemically (i.p.), or mucosally (i.e., intranasally or intratracheally). Interestingly, a single dose of ISS inhibited airway eosinophilia as effectively as daily injections of corticosteroids for 7 days. Moreover, while both ISS and corticosteroids inhibited IL-5 generation, only ISS was able to induce allergen-specific IFN-gamma production and redirect the immune system toward a Th1 response. Thus, systemic or mucosal administration of ISS before allergen exposure could provide a novel form of active immunotherapy in allergic diseases.

Adjuvants, Immunologic↗
PubMed1998

The crystal structure of the IkappaBalpha/NF-kappaB complex reveals mechanisms of NF-kappaB inactivation.

IkappaBalpha regulates the transcription factor NF-kappaB through the formation of stable IkappaBalpha/NF-kappaB complexes. Prior to induction, IkappaBalpha retains NF-kappaB in the cytoplasm until the NF-kappaB activation signal is received. After activation, NF-kappaB is removed from gene promoters through association with nuclear IkappaBalpha, restoring the preinduction state. The 2.3 A crystal structure of IkappaBalpha in complex with the NF-kappaB p50/p65 heterodimer reveals mechanisms of these inhibitory activities. The presence of IkappaBalpha allows large en bloc movement of the NF-kappaB p65 subunit amino-terminal domain. This conformational change induces allosteric inhibition of NF-kappaB DNA binding. Amino acid residues immediately preceding the nuclear localization signals of both NF-kappaB p50 and p65 subunits are tethered to the IkappaBalpha amino-terminal ankyrin repeats, impeding NF-kappaB from nuclear import machinery recognition.

Amino Acid Sequence↗
PubMed1998

Ikappa Balpha functions through direct contacts with the nuclear localization signals and the DNA binding sequences of NF-kappaB.

We have determined the binding energies of complexes formed between Ikappa Balpha and the wild type and mutational variants of three different Rel/NF-kappaB dimers, namely, the p50/p65 heterodimer and homodimers of p50 and p65. We show that although a common mode of interaction exists between the Rel/NF-kappaB dimers and Ikappa Balpha, IkappaB alpha binds the NF-kappaB p50/p65 heterodimer with 60- and 27-fold higher affinity than the p50 and p65 homodimers, respectively. Each of the three flexibly linked segments of the rel homology region of Rel/NF-kappaB proteins (the nuclear localization sequence, the dimerization domain, and the amino-terminal DNA binding domain) is directly engaged in forming the protein/protein interface with the ankyrin repeats and the carboxyl-terminal acidic tail/PEST sequence of Ikappa Balpha. In the cell, Ikappa Balpha functions to retain NF-kappaB in the cytoplasm and inhibit its DNA binding activity. These properties are a result of the direct involvement of the nuclear localization sequences and of the DNA binding region of NF-kappaB in complex with Ikappa Balpha. A model of the interactions in the complex is proposed based on our observations and the crystal structures of Rel/NF-kappaB dimers and the ankyrin domains of related proteins.

Base Sequence↗
PubMed1998

[Sinus node dysfunction after complete correction of tetralogy of Fallot. Report of a case].

Atrioventricular conduction defects after complete correction of tetralogy of Fallot are well known and their prevalence is decreasing with improved operative techniques. On the other hand, sinus node dysfunction has not been described after this type of surgery. The authors report the case of a 24 year old patient operated at the age of 2 for a favourable form of tetralogy of Fallot. In the long-term, this patient had a minimal anatomical sequel (mild pulmonary insufficiency) and first degree AVB with complete right bundle branch block with sinus bradycardia on the surface ECG. A recent aggravation of the patient's functional status led to rhythmological investigation which showed chronotropic atrial incompetence with nodal atrioventricular delay. The patient was implanted with a permanent DDD cardiac pacemaker programmed in the DDDR mode with a good outcome. Atrial chronotropic incompetence is one of the modes of expression of sinus node dysfunction. This has not been previously described in this context. This form of sinus node dysfunction should be systematically excluded after surgical cure of tetralogy of Fallot by exercise stress testing and 24 hour Holter ECG. Symptomatic patients are greatly improved by permanent cardiac pacing with hysteresis.

Adult↗
PubMed1997

A prospective study of the effect of I.V. pentamidine therapy on ventricular arrhythmias and QTc prolongation in HIV-infected patients.

STUDY OBJECTIVES: I.V. pentamidine therapy in HIV-infected patients has been associated in case reports and one uncontrolled prospective series with frequent prolongation of the rate-corrected QT interval (QTc) and a high risk for potentially lethal ventricular arrhythmias, especially torsade de pointes. The aim of this study was to prospectively examine in a controlled manner the effect of I.V. pentamidine therapy on the QT interval and the incidence of ventricular arrhythmias. DESIGN: Open, nonrandomized, prospective evaluation of ventricular arrhythmia incidence in HIV-infected patients receiving pentamidine or trimethoprim-sulfamethoxazole (TMP-SMX) utilizing Holter monitoring prior to and during therapy with these agents. SETTING: Staten Island University Hospital, Staten Island, NY. PATIENTS: Twenty-seven HIV-infected patients, of whom 16 received I.V. pentamidine and 11 received I.V. TMP-SMX. MEASUREMENTS AND RESULTS: Study patients underwent Holter monitoring prior to therapy and during the first 3 days and last 2 days of therapy with pentamidine or TMP-SMX, 12-lead ECG prior to and every 24 to 48 h, serum electrolytes prior to and on days 3, 6, 9, and 12 of therapy, and baseline transthoracic two-dimensional and Doppler echocardiography. In the pentamidine group, the results for each monitoring period were as follows (means are presented +/- SEM): pretherapy, 1.66+/-1.03 (median=0) premature ventricular complexes (PVCs) per hour, zero nonsustained ventricular tachycardia (NSVT), zero sustained ventricular tachycardia (VT); early therapy, 1.55+/-0.91 (median=0.04) PVCs per hour, two NSVT (both < or = 5 complexes), zero sustained VT; late therapy, 1.69+/-1.17 (median=0.08) PVCs per hour, zero NSVT, zero sustained VT (p value not significant for early or late therapy as compared to pretherapy for PVCs per hour, NSVT, or sustained VT). In the TMP-SMX group, the Holter monitoring results were as follows: pretherapy, 1.36+/-1.27 (median=0) PVCs per hour, zero NSVT, zero sustained VT; early therapy, 0.71+/-0.53 (median=0.03) PVCs per hour, two NSVT, zero sustained VT; late therapy, 0.56+/-0.51 (median=0) PVCs per hour, zero NSVT, zero sustained VT (p value not significant for pretherapy, early therapy, or late therapy with TMP-SMX as compared to pentamidine for PVCs per hour, NSVT, or VT). The QTc also did not significantly differ during therapy with pentamidine as compared to TMP-SMX. The mean QTc in the pentamidine group decreased during therapy as compared to pretherapy with the difference approaching significance for days 2, 4, and 6 with pentamidine (p<0.06). CONCLUSIONS: QTc prolongation during therapy with pentamidine in HIV-infected patients is not as frequent an occurrence as has been reported previously. In the absence of QTc prolongation, pentamidine therapy was not associated with a significant increase in PVCs, NSVT, or sustained VT as compared to pretherapy recordings or as compared to therapy with TMP-SMX.

Adult↗
PubMed1995

The status of local smoking regulations in North Carolina following a state preemption bill.

OBJECTIVE: To determine the number and protectiveness of local smoking regulations adopted before the implementation of a preemptive statewide smoking control bill. METHOD: Review of local smoking control regulations from all 100 counties and 85 municipalities with populations greater than 5000 in North Carolina. MAIN OUTCOME MEASURES: Adoption of local smoking control regulations before and during the 3-month delay in enactment of the preemptive bill. Protectiveness of regulations was based on restrictions on smoking and requirements for separate ventilation systems at private work sites: none (smoking unrestricted); minimal (smoking restricted to designated areas); partial (smoking restricted to designated areas served by separate ventilation systems); and complete (smoking prohibited). Because some regulations would be phased in gradually over the next 5 years, we evaluated the requirements that will be in effect by January 1, 2000. RESULTS: Between July 15 and October 15, 1993, the number of local smoking regulations in North Carolina increased from 16 to 105. By the year 2000, 59% of private employees still will not be guaranteed any protection from work site environmental tobacco smoke; 19% will have minimal protection, 22% will have partial protection, and none will have complete protection. CONCLUSIONS: The 3-month delay in preemption created an unnatural time frame for communities to organize, debate, and adopt smoking restrictions. Despite the adoption of 89 new regulations, no private employees will be guaranteed complete protection from work site environmental tobacco smoke by the year 2000; new regulations can no longer be adopted. HB 957 has been a setback for public health in North Carolina.

Humans↗
PubMed1992

Structure of a G.T.A triplet in an intramolecular DNA triplex.

A 32-base DNA oligonucleotide has been studied by one- and two-dimensional 1H NMR spectroscopy and is shown to form a stable, pyr.pur.pyr, intramolecular triple helical structure, with a four C loop and a TATA loop connecting the Watson-Crick- and Hoogsteen-paired strands, respectively. This triplex contains five T.A.T base triplets, two C+.G.C base triplets, and an unusual G.T.A base triplet which disrupts the pyr.pur.pyr motif. The G.T.A triplet consists of a Watson-Crick T.A base pair, with the T situated in the "purine strand" and the A situated in the "pyrimidine strand" and a G situated in the Hoogsteen-base-paired "pyrimidine strand" hydrogen bonded to the T. The base-pairing structure of the G.T.A triplet has been investigated and has been found to involve a single hydrogen bond from the guanine amino group to the O4 carbonyl of the thymine, leaving the guanine imino proton free. The specific amino proton involved in the hydrogen bond is the H2(2) proton. This orients the guanine such that its sugar is near the thymine methyl group. The guanine sugar adopts an N-type (C3'-endo) sugar pucker in this triplet. The stability of the G.T.A triplet within pyr.pur.pyr triplexes is discussed.

Base Composition↗
PubMed1991

Structure and stability of X.G.C mismatches in the third strand of intramolecular triplexes.

Intramolecular DNA triplexes that contain eight base triplets formed from the folding of a single DNA strand tolerate a single X.G.C mismatch in the third strand at acidic pH. The structure and relative stability of all four triplets that are possible involving a G.C Watson-Crick base pair were determined with one- and two-dimensional proton nuclear magnetic resonance techniques. Triplexes containing A.G.C, G.G.C, or T.G.C triplets were less stable than the corresponding parent molecule containing a C.G.C triplet. However, all mismatched bases formed specific hydrogen bonds in the major groove of the double helix. The relative effect of these mismatches on the stability of the triplex differs from the effect assayed (under different conditions) by two-dimensional gel electrophoresis and DNA cleavage with oligonucleotide EDTA.Fe(II).

Base Sequence↗
PubMedDate not supplied

Early organ-specific hemorrhage-induced increases in tissue cytokine content: associated neurohormonal and opioid alterations.

Hemorrhage is associated with an impairment in the immune response and with increased concentrations of circulating inflammatory cytokines. The present study determined the time course and localization of alterations in circulating and tissue pro-inflammatory cytokines (TNF-alpha, IL-1-alpha and -beta) in response to fixed-pressure (40 mm Hg) hemorrhage as well as the associated hanges in circulating neurohormonal and opioid mediators. Conscious unrestrained non-heparinized male Sprague-Dawley rats (n = 24) underwent hemorrhage followed by standard resuscitation with lactated Ringer's solution. Animals were sacrificed at three time points; immediately after the hemorrhage period, at completion of resuscitation and 1.5 h after the resuscitation period. Hemorrhage resulted in marked elevations in circulating levels of TNF-alpha, which averaged 860 +/- 201 pg/ml. The levels were similarly elevated following fluid resuscitation (877 +/- 196 pg/ml) and had decreased towards baseline 1.5 h after completion of resuscitation (281 +/- 134 pg/ml). TNF-alpha was not detectable in plasma of time-matched controls. Hemorrhage elevated TNF-alpha content in spleen (25%), lung (55%) and heart (20%), and tissue content remained elevated despite resuscitation. No significant changes in tissue content of TNF-alpha were detected in the liver, kidney or brain. Circulating levels of IL1-alpha and -beta were not detectable in either the time-matched controls or hemorrhaged animals. However, statistically significant elevations in tissue content of IL-1 alpha were observed in heart, spleen, lung, gut and whole brain (15-30%). Tissue content of IL-1 beta did not change in response to hemorrhage and/or fluid resuscitation. Activation of sympathetic outflow, as evidenced by a 3- to 4-fold elevation in circulating epinephrine and norepinephrine levels, was observed immediately after hemorrhage, and was associated with a 5-fold rise in circulating beta-endorphin. These results demonstrate an early increase in tissue cytokine content following hemorrhagic shock, which is associated with elevations in circulating catecholamines and endogenous opioids, consistent with their potential modulatory role in this response.

Animals↗