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71.
72.
IS Efimova A Salminen P Pohjanjoki J Lapinniemi NN Magretova BS Cooperman A Goldman R Lahti AA Baykov 《Canadian Metallurgical Quarterly》1999,274(6):3294-3299
Recent crystallographic studies on Escherichia coli inorganic pyrophosphatase (E-PPase) have identified three Mg2+ ions/enzyme hexamer in water-filled cavities formed by Asn24, Ala25, and Asp26 at the trimer-trimer interface (Kankare, J., Salminen, T., Lahti, R., Cooperman, B., Baykov, A. A., and Goldman, A. (1996) Biochemistry 35, 4670-4677). Here we show that D26S and D26N substitutions decrease the stoichiometry of tight Mg2+ binding to E-PPase by approximately 0.5 mol/mol monomer and increase hexamer stability in acidic medium. Mg2+ markedly decelerates the dissociation of enzyme hexamer into trimers at pH 5.0 and accelerates hexamer formation from trimers at pH 7.2 with wild type E-PPase and the N24D variant, in contrast to the D26S and D26N variants, when little or no effect is seen. The catalytic parameters describing the dependences of enzyme activity on substrate and Mg2+ concentrations are of the same magnitude for wild type E-PPase and the three variants. The affinity of the intertrimer site for Mg2+ at pH 7.2 is intermediate between those of two Mg2+ binding sites found in the E-PPase active site. It is concluded that the metal ion binding site found at the trimer-trimer interface of E-PPase is a high affinity site whose occupancy by Mg2+ greatly stabilizes the enzyme hexamer but has little effect on catalysis. 相似文献
73.
Iron superoxide dismutase from the archaeon Sulfolobus solfataricus: analysis of structure and thermostability 总被引:1,自引:0,他引:1
T Ursby BS Adinolfi S Al-Karadaghi E De Vendittis V Bocchini 《Canadian Metallurgical Quarterly》1999,286(1):189-205
The crystal structure of superoxide dismutase (SOD) from the hyper thermophile Sulfolobus solfataricus has been determined at 2.3 A resolution by molecular replacement and refined to a crystallographic R-factor of 16.8 % (Rfree 19.8 %). The crystals belong to the space group C2 (a=76.3 A, b=124.3 A, c=60.3 A, beta=128.8 degrees) with two identical monomers in the asymmetric unit. The monomer has a molecular weight of 24 kDa and consists of 210 amino acid residues of which 205 are visible in the electron density map. The overall fold of the monomer of S. solfataricus SOD is similar to that of the other known Fe or Mn-SODs. S. solfataricus SOD forms a very compact tetramer of a type similar to that of SOD from the hyperthermophile Aquifex pyrophilus. Both structures show an elevated number of inter-subunit ion-pairs compared with the mesophilic SOD from Mycobacterium tuberculosis and the thermophilic SOD from Thermus thermophilus. However, in contrast to the A. pyrophilus SOD structure, the number of intra-subunit ion-pairs as well as inter- subunit hydrogen bonds is not higher than in the compared mesophilic and thermophilic SOD structures. The electron density also revealed an unexpected and unusual covalent modification of a conserved tyrosine in the active site. Its involvement in the specific activity of the enzyme is discussed. 相似文献
74.
BS Jassar KH Harris PM Ostashewski JH Jhamandas 《Canadian Metallurgical Quarterly》1999,81(1):234-246
Whole cell recordings were performed on acutely dissociated neurons from the horizontal limb of the diagonal band of Broca (hDBB) from rats to elucidate the ionic mechanisms of action of neurotensin. Neurotensin caused a decrease in whole cell voltage-activated outward currents and failed to elicit a response when Ca2+ influx was blocked by changing the external solution to the one containing 0 mM Ca2+ and 50 microM Cd2+, suggesting the involvement of Ca2+-dependent conductances. Charybdotoxin, a specific blocker of voltage-sensitive calcium-activated K+ channels (IC), caused a decrease in outward currents comparable with that caused by blocking calcium influx and occluded the neurotensin-induced decrease in outward currents. Similarly, 50 microM tetraethylammonium ions also blocked the neurotensin response. Also neurotensin reduced whole cell barium currents (IBa) and calcium currents (ICa). Amiloride and omega-conotoxin GVIA, but not nimodipine, were able to eliminate the neurotensin-induced decrease in IBa. Thus T- and N- but not L-type calcium channels are subject to modulation by neurotensin, and this may account for its effects on IC. The predicted changes in action potential as a result of the blockade of currents through calcium channels culminating into changes in IC were confirmed in the bridge current-clamp recordings. Specifically, neurotensin application led to depolarization of the resting membrane potential, broadening of spike and a decrease in afterhyperpolarization and accommodation. These alterations in action potential characteristics that resulted in increased firing rate and excitability of the hDBB neurons also were produced by application of charybdotoxin. Neurotensin effects on these properties were occluded by 2 - [(1 - 7 - chloro - 4 - quinolinyl) - 5 - (2, 6 - di - methoxyphenyl) pyrazol-3-yl) carbonylamino] tricyclo (3.3.1.1.)decan-2-carboxylic acid, a nonpeptide high-affinity neurotensin receptor antagonist. Neurotensin blockade of IC, possibly through ICa, is a potential physiological mechanism whereby this peptide may evoke alterations in the cortical arousal, sleep-wake cycle, and theta rhythm. 相似文献
75.
A. N. Gruzintsev G. A. Emelchenko A. N. Redkin W. T. Volkov E. E. Yakimov G. Visimberga S. G. Romanov 《Semiconductors》2009,43(4):468-471
The scanning cathodoluminescence spectra and images of a ZnO nanorod with two mirrors at the end faces are studied at 10 K. Emission peaks of bound excitons and edge luminescence are observed in the ultraviolet spectral region. The spatial distribution pattern of emission intensity corresponding to localized photon modes of the nanorod is recorded at different wavelengths. A new method of visualization of localized optical modes in micro- and nano-sized laser cavities is suggested. 相似文献
76.
Vincristine was inadvertently injected into a thigh of three children. In each case the accident occurred as a result of the mixing of a syringe containing vincristine with a syringe of L-asparaginase which the patient was scheduled to receive on the same day. Within minutes, each patient was treated topically with cold compresses and the area was infiltrated with a solution of 8.4% sodium bicarbonate. Only one patient had discomfort of the thigh after the injection, none of the patients have had any sequelae, either acute or delayed. Measures to avoid mistaken injection of vincristine for asparaginase are readily achievable and have prevented recurrences of intramuscular vincristine administration at the institutions where they have been implemented. Nonetheless, other instances of intramuscular vincristine injection are anticipated and should be rapidly recognized and quickly managed with local applications of cold and sodium bicarbonate. 相似文献
77.
78.
79.
The convective transport of fluid and of a binding antibody through a cultured tumor cell layer was investigated with a mouse melanoma cell line (B16F10) grown on a microporous polycarbonate filter (Snapwell inserts). The inserts were precoated with Matrigel or collagen, or were uncoated. The cell layers were exposed to nominal pressure gradients from 5 to 25 cm H2O, and the volume flux was measured by collecting the effluent volume over time. The rate of convective transport of a binding monoclonal antibody that recognizes the murina transferrin receptor (a-TfR) was investigated at a nominal pressure gradient of 15 cm H2O and compared with that of an isotype matched, nonbinding control. The resistance, R, of the cell layer to fluid flow was quantified as the hydraulic conductivity, Lp (= 1/R); the ability of the cell layer to retard antibody transport was quantified as the reflection coefficient, sigma. The resulting Lp values decreased with increasing cell density, in a manner consistent with Poiseuille flow. Collagen or Matrigel precoating also decreased Lp values, with cells grown on Matrigel providing the greatest resistance. The sigma values were 0.67 (+/-0.08) for the a-TfR antibody and 0.51 (+/-0.06) for the control, indicating that the cell layer acts as a semipermeable barrier to convective transport of antibody that is less permeable to the binding antibody. 相似文献
80.
BD Spiess MH Wall BS Gillies JC Fitch LO Soltow WL Chandler 《Canadian Metallurgical Quarterly》1997,78(2):820-826
Thromboelastography (TEG) has been used after cardiopulmonary bypass (CPB) to diagnose excessive postoperative hemorrhage. Conventional TEG during CPB is not possible due to the sensitivity of the TEG to even small amounts of heparin, which produces a nondiagnostic tracing. The purpose of this study was to compare heparin neutralization using heparinase or protamine in TEG blood samples obtained during CPB. TEG testing was performed on 48 patients before, during and after CPB. Tissue plasminogen activator activity and antigen were measured on a subset of 32 patients. We found: 1) heparinase neutralized at least 10 IU/ml heparin while 1.6 ug/ml protamine neutralized up to 7 IU/ml heparin, 2) in samples with complete heparin neutralization by both methods, there was no significant difference in the R values, 3) while there was good correlation for other TEG parameters between heparinase and protamine treated samples, heparinase treatment produced shorter K values and higher angle, MA and A60, 4) while fibrinolysis was detected using both methods, heparinase treatment suppressed fibrinolysis in the TEG in both samples from patients and after in vitro addition of tissue plasminogen activator, 5) TEG was not a sensitive indicator of t-PA activity, detecting only 21% of samples with increased t-PA activity during bypass, and 5) heparinase was at least 100 times more expensive than protamine. We conclude that while both heparinase and protamine can be used to neutralize heparin in TEG samples obtained during CPB, protamine neutralization is more sensitive to fibrinolysis and less expensive, but the protamine dose must be carefully selected to match the heparin level used at individual institutions. 相似文献