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61.
Growth of Aeropyrum pernix, the first reported aerobic neutrophilic hyperthermophilic archaeon, was investigated under different cultivation parameters. Different sources of seawater, pH, and the cultivation methods were tested with the aim to improve the biomass production. A 1-L glass flask fitted with a condenser and air diffuser was used as a bioreactor. The optimum conditions for maximizing A. pernix biomass were obtained when Na2S2O3.5H2O (1 g/L) with added marine broth 2216 at pH 7.0 (20 mmol HEPES buffer/L) was used as a growing medium in a 1-L flask. The biomass production was 0.45 g dry cell mass/L in 40 h under the optimum conditions, which is more than the 0.42 g dry cell mass/L in 60 h previously obtained. 相似文献
62.
Roumenina LT Kantardjiev AA Atanasov BP Waters P Gadjeva M Reid KB Mantovani A Kishore U Kojouharova MS 《Biochemistry》2005,44(43):14097-14109
C1q is the recognition subunit of the classical pathway of the complement system and a major connecting link between classical pathway-driven innate immunity and IgG- or IgM-mediated acquired immunity. The basic structural subunit of C1q is composed of an N-terminal triple-helical collagen-like region and a C-terminal heterotrimeric globular head domain (gC1q) that is made up of individual A, B, and C chains. Recent crystallographic studies have revealed that the gC1q domain, which is the main target-binding region of C1q, has a compact and spherical heterotrimeric assembly, held together by both electrostatic and nonpolar interactions, with quasi-3-fold symmetry. A characteristic feature of the gC1q domain is the presence of a exposed Ca(2+) located near the apex. We have investigated, using theoretical and experimental approaches, the role of Ca(2+) in the electrostatic stability and target-binding properties of the native C1q as well as recombinant monomeric forms of the C-terminal regions of the A, B, and C chains. Here, we report that Ca(2+) primarily influences the target recognition properties of C1q toward IgG, IgM, C-reactive protein, and pentraxin 3. At pH 7.4, the loss of Ca(2+) leads to changes in the direction of electric moment from coaxial (where the putative C-reactive protein-binding site is located) to perpendicular to the molecular axis (toward the most likely IgG-binding site), which appears important for target recognition by C1q and subsequent complement activation. 相似文献
63.
Alzheimer's disease is defined in part by the intraneuronal aggregation of tau protein into filamentous lesions. The pathway is accompanied by posttranslational modifications including phosphorylation and glycation, each of which has been shown to promote tau fibrillization in vitro when present at high stoichiometry. To clarify the site-specific impact of posttranslational modification on tau fibrillization, the ability of recombinant full-length four repeat tau protein (htau40) and 11 pseudophosphorylation mutants to fibrillize in the presence of anionic inducer was assayed in vitro using transmission electron microscopy and laser light scattering assays. Tau glycated with d-glucose was examined as well. Both glycated tau and pseudophosphorylation mutants S199E, T212E, S214E, double mutant T212E/S214E, and triple mutant S199E/S202E/T205E yielded increased filament mass at equilibrium relative to wild-type tau. Increases in filament mass correlated strongly with decreases in critical concentration, indicating that both pseudophosphorylation and glycation promoted fibrillization by shifting equilibrium toward the fibrillized state. Analysis of reaction time courses further revealed that increases in filament mass were not associated with reduced lag times, indicating that these posttranslational modifications did not promote filament nucleation. The results suggest that site-specific posttranslational modifications can stabilize filaments once they nucleate, and thereby support their accumulation at low intracellular tau concentrations. 相似文献
64.
Static light scattering is an important solution-based method for assaying spontaneous protein aggregation reactions. But the reliability of the measurements when conducted in the presence of fibrillization inducers has been questioned. Here the utility of static laser light scattering for quantitative assay of anionic micelle-induced protein fibrillization was characterized using tau protein, the major component of neurofibrillary lesions of Alzheimer's disease. Both inducer micellization and tau fibrillization made significant contributions to light scattering intensity. The intensity arising solely from micellization was quantified using proteins that promoted inducer micellization but could not fibrillize, such as mixed histones and assembly-incompetent mutant htau40(I277P/I308P). When corrected for micellization, reaction progress curves for wild-type tau fibrillization were sigmoidal and correlated well with measurements of total filament length made by transmission electron microscopy. The utility of the improved laser light scattering assay was demonstrated by quantifying the effect of inducer concentration on tau assembly kinetics using a three-parameter Gompertz growth function. Results showed that alkyl sulfate detergent accelerated tau nucleation as reflected by shorter lag times and modulated pre-nuclear equilibria to yield more filament mass at reaction equilibrium. 相似文献
65.
Kojouharova MS Gadjeva MG Tsacheva IG Zlatarova A Roumenina LT Tchorbadjieva MI Atanasov BP Waters P Urban BC Sim RB Reid KB Kishore U 《Journal of immunology (Baltimore, Md. : 1950)》2004,172(7):4351-4358
The first step in the activation of the classical complement pathway by immune complexes involves the binding of the globular domain (gC1q) of C1q to the Fc regions of aggregated IgG or IgM. Each gC1q domain is a heterotrimer of the C-terminal halves of one A (ghA), one B (ghB), and one C (ghC) chain. Our recent studies have suggested a modular organization of gC1q, consistent with the view that ghA, ghB, and ghC are functionally autonomous modules and have distinct and differential ligand-binding properties. Although C1q binding sites on IgG have been previously identified, the complementary interacting sites on the gC1q domain have not been precisely defined. The availability of the recombinant constructs expressing ghA, ghB, and ghC has allowed us, for the first time, to engineer single-residue substitution mutations and identify residues on the gC1q domain, which are involved in the interaction between C1q and IgG. Because C1q is a charge pattern recognition molecule, we have sequentially targeted arginine and histidine residues in each chain. Consistent with previous chemical modification studies and the recent crystal structure of gC1q, our results support a central role for arginine and histidine residues, especially Arg(114) and Arg(129) of the ghB module, in the C1q-IgG interaction. 相似文献
66.
Pavlicev M Mayer W 《Journal of experimental zoology. Part B. Molecular and developmental evolution》2006,306(6):539-550
The analysis of a 581 bp section of the nuclear gene c-mos revealed multiple copies of putative functional sequences as well as pseudogenes in three closely related lacertid species Lacerta laevis, L. kulzeri and L. cyanisparsa. A phylogenetic analysis of c-mos in comparison with a molecular phylogeny based on the mitochondrial cytochrome b gene supports our findings. The study also provides new insights into the phylogenetic relationships of L. cyanisparsa and L. laevis.Pseudogenes of the three species share 11 single-nucleotide substitutions, a 1 bp deletion and a premature stop codon but differ by group-specific mutations. This result suggests that the c-mos gene has become duplicated and subsequently silenced already in the common ancestor of the three species. Sequence divergence suggests that the duplication and the loss of function occurred in the late Miocene/early Pliocene, i.e., about 5 million years ago. Indications of gene conversion are discussed.We suggest that future studies using c-mos for phylogenetic studies should provide evidence for the orthology of the sequences compared. 相似文献
67.
Hinescu ME Gherghiceanu M Mandache E Ciontea SM Popescu LM 《Journal of cellular and molecular medicine》2006,10(1):243-257
We have previously reported (Hinescu & Popescu, 2005) the existence of interstitial Cajal-like cells (ICLC), by transmission electron microscopy, in human atrial myocardium. In the present study, ICLC were identified with non-conventional light microscopy (NCLM) on semi-thin sections stained with toluidine blue and immunohistochemistry (IHC) for CD117/c-kit, CD34, vimentin and other additional antigens for differential diagnosis. Quantitatively, on semi-thin sections, ICLC represent about 1-1.5% of the atrial myocardial volume (vs. approximately 45% working myocytes, approximately 2% endothelial cells, 3-4% for other interstitial cells, and the remaining percentage: extracellular matrix). Roughly, there is one ICLC for 8-10 working atrial myocytes in the intercellular space, beneath the epicardium, with a characteristic (pyriform, spindle or triangular) shape. These ICLC usually have 2-3 definitory processes, emerging from cell body, which usually embrace atrial myocytes (260 nm average distance plasmalemma/sarcolemma) or establish close contact with nerve fibers or capillaries (approximately 420 nm average distance to endothelial cells). Cell prolongations are characteristic: very thin (mean thickness = 0.15+/-0.1 microm), very long for a non-nervous cell (several tens of microm) and moniliform (uneven caliber). Stromal synapses between ICLC and other interstitial cells (macrophages) were found (e.g. in a multicontact type synapse, the average synaptic cleft was approximately 65 nm). Naturally, the usual cell organelles (mitochondria, smooth and rough endoplasmic reticulum, intermediate filaments) are relatively well developed. Caveolae were also visible on cell prolongations. No thick filaments were detected. IHC showed that ICLC were slightly and inconsistently positive for CD117/c-kit, variously co-expressed CD34 and EGF receptor, but appeared strongly positive for vimentin, along their prolongations. Some ICLC seemed positive for a-smooth muscle actin and tau protein, but were negative for nestin, desmin, CD13 and S-100. In conclusion, we provide further evidence of the existence of ICLC in human atrial myocardium, supporting the possible ICLC role in pacemaking, secretion (juxta- and/or paracrine), intercellular signaling (neurons and myocytes). For pathology, ICLC might as well be 'players' in arrhythmogenesis and atrial remodeling. 相似文献
68.
Hutterer A Berdnik D Wirtz-Peitz F Zigman M Schleiffer A Knoblich JA 《Developmental cell》2006,11(2):147-157
The protein kinase Aurora-A is required for centrosome maturation, spindle assembly, and asymmetric protein localization during mitosis. Here, we describe the identification of Bora, a conserved protein that is required for the activation of Aurora-A at the onset of mitosis. In the Drosophila peripheral nervous system, bora mutants have defects during asymmetric cell division identical to those observed in aurora-A. Furthermore, overexpression of bora can rescue defects caused by mutations in aurora-A. Bora is conserved in vertebrates, and both Drosophila and human Bora can bind to Aurora-A and activate the kinase in vitro. In interphase cells, Bora is a nuclear protein, but upon entry into mitosis, Bora is excluded from the nucleus and translocates into the cytoplasm in a Cdc2-dependent manner. We propose a model in which activation of Cdc2 initiates the release of Bora into the cytoplasm where it can bind and activate Aurora-A. 相似文献
69.
Yang YH Hwang KK FitzGerald J Grossman JM Taylor M Hahn BH Chen PP 《Journal of immunology (Baltimore, Md. : 1950)》2006,177(11):8219-8225
Antiphospholipid Ab have been shown to promote thrombosis and fetal loss in the antiphospholipid syndrome (APS). Previously, we found IgG anti-thrombin Ab in some APS patients that could interfere with inactivation of thrombin by antithrombin (AT). Considering that activated coagulation factor X (FXa) is homologous to thrombin in the catalytic domains and is also regulated primarily by AT, we hypothesized that some thrombin-reactive Ab may bind to FXa and interfere with AT inactivation of FXa. To test these hypotheses, we studied reactivity of eight patient-derived monoclonal IgG antiphospholipid Ab with FXa and the presence of IgG anti-FXa Ab in APS patients and investigated the effects of FXa-reactive mAb on AT inactivation of FXa. The results revealed that six of six thrombin-reactive IgG mAb bound to FXa and that the levels of plasma IgG anti-FXa Ab in 38 APS patients were significantly higher than those in 30 normal controls (p < 0.001). When the mean plus 3 SDs of the 30 normal controls was used as the cutoff, 5 of 38 APS patients (13.2%) had IgG anti-FXa Ab. Importantly, three of six FXa-reactive mAb significantly inhibited AT inactivation of FXa. Combined, these results indicate that anti-FXa Ab may contribute to thrombosis by interfering with the anticoagulant function of AT on FXa in some APS patients. 相似文献
70.
The Structure of Polyunsaturated Lipid Bilayers Important for Rhodopsin Function: A Neutron Diffraction Study
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Mihaela Mihailescu 《Biophysical journal》2006,90(1):L04
The structure of oriented 1-stearoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine bilayers with perdeuterated stearoyl- or docosahexaenoyl hydrocarbon chains was investigated by neutron diffraction. Experiments were conducted at two different relative humidities, 66 and 86%. At both humidities we observed that the polyunsaturated docosahexaenoyl chain has a preference to reside near the lipid water interface. That leaves voids in the bilayer center that are occupied by saturated stearoyl chain segments. This uneven distribution of saturated- and polyunsaturated chain densities is likely to result in membrane elastic stress that modulates function of integral receptor proteins like rhodopsin. 相似文献