A Mutational Analysis of DNA Binding and Cleavage by the Homing Endonuclease PI-SceI

We have carried out an extensive mutational analysis of PI-SceI, the best studied intein-like homing endonuclease of the LAGLIDADG family, to find out which amino acid residues are involved in substrate binding and processing. Our analysis was focused on domain I, in which two regions were shown to be in contact with DNA, and on domain II, in which the amino acid residues making up catalytic centers I and II were identified and their role in catalysis investigated. As a result of our comprehensive mutational analysis a model is presented for DNA binding and cleavage by PI-SceI.     

Data management for large‐scale scientific computations in high performance distributed systems

With the increasing number of scientific applications manipulating huge amounts of data, effective high-level data management is an increasingly important problem. Unfortunately, so far the solutions to the high‐level data management problem either require deep understanding of specific storage architectures and file layouts (as in high-performance file storage systems) or produce unsatisfactory I/O performance in exchange for ease-of-use and portability (as in relational DBMSs). In this paper we present a novel application development environment which is built around an active meta-data management system (MDMS) to handle high-level data in an effective manner. The key components of our three-tiered architecture are user application, the MDMS, and a hierarchical storage system (HSS). Our environment overcomes the performance problems of pure database-oriented solutions, while maintaining their advantages in terms of ease-of-use and portability. The high levels of performance are achieved by the MDMS, with the aid of user-specified, performance-oriented directives. Our environment supports a simple, easy-to-use yet powerful user interface, leaving the task of choosing appropriate I/O techniques for the application at hand to the MDMS. We discuss the importance of an active MDMS and show how the three components of our environment, namely the application, the MDMS, and the HSS, fit together. We also report performance numbers from our ongoing implementation and illustrate that significant improvements are made possible without undue programming effort. This revised version was published online in July 2006 with corrections to the Cover Date.     

The effects of liposome-reconstituted apolipoproteins on the binding of rat intermediate density lipoproteins to rat liver membranes

Rat intermediate density lipoproteins (IDL) bind specifically to high and low affinity binding sites on rat liver membranes. In a recent paper (Brissette, L., and No?l, S.-P. (1986) J. Biol. Chem. 261, 6847-6852), we have demonstrated that human low density lipoproteins and high density lipoproteins-3 can totally prevent the specific binding of rat IDL to the low affinity binding sites. The aim of the present studies was to determine the effects of apoA-I, apoC, and apoE, reconstituted into liposomes, on the binding of rat iodinated IDL to rat liver membranes. We found that a 50-, 100-, or 300-fold excess of liposome-reconstituted apoE, apoC, or apoA-I, respectively, abolished the specific binding of IDL to the low affinity binding sites. Only apoE liposomes had an effect on the high affinity component; at a 100-fold excess no specific binding of IDL could be detected. Liposomes by themselves or associated with erythrocyte membrane proteins had virtually no effect on the binding of IDL. Taken together our results suggest that apoE is the only ligand that can compete efficiently for the sites that bind rat IDL with a high affinity. These sites may be the expression of both the remnant and the LDL receptors. The binding to the low affinity component probably represents weak interactions between IDL and "unspecified-lipoprotein binding sites," which can be entirely masked by human low density lipoproteins, high density lipoproteins-3, or liposome-reconstituted apoA-I, apoE, or apoC at appropriate concentrations.     

Tissue calmodulin levels in normal and Graves' thyroids

Calmodulin levels in normal human thyroids and Graves' disease thyroids were measured by specific radioimmunoassay in the presence of ethyleneglycol-bis-(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). The calmodulin levels in tissues from patients with Graves' disease treated with thionamide drugs were significantly higher than those in normal tissues from euthyroid patients with solitary cold nodules (normal: 484 +/- 50 ng/mg protein, mean +/- SE, n = 15; Graves': 901 +/- 54 ng/mg protein, n = 48, p less than 0.001). Such a rise in calmodulin levels in Graves' disease thyroids was also present even after the administration of 50 micrograms of T3 for 5 days before operation (828 +/- 137 ng/mg protein, n = 6, p less than 0.01). Calmodulin levels in Graves' disease thyroids were closely related to the cell height of follicular epithelium. Calmodulin levels in a columnar cell predominant group were significantly higher than those in a flat cell predominant or a cuboidal cell predominant group (columnar cell predominant: 1150 +/- 118 ng/mg protein, n = 13; flat cell predominant: 561 +/- 125 ng/mg protein, n = 3, p less than 0.05; cuboidal cell predominant: 596 +/- 40 ng/mg protein, n = 25, p less than 0.001). The increase in calmodulin content in Graves' disease thyroid could therefore possibly be attributed to the stimulation of the thyroid gland by the thyroid stimulating antibody. An immunofluorescence study demonstrated the presence of calmodulin immunoreactivity in the thyroid epithelial cells, particularly enriched in the apical border in the form of a granulated structure.     

In vitro synthesis and assembly of a 68 kDa outer mitochondrial membrane protein from rat liver

Outer mitochondrial membrane was purified from rat liver. Its constituent proteins were analyzed by SDS-polyacrylamide gel electrophoresis and by electrophoretic immunoblotting employing antibodies raised against total outer mitochondrial membrane. Anti-outer mitochondrial membrane antiserum reacted with only one polypeptide (15 kDa) in rough microsomes, whereas no immunological cross-reactivity was observed with other mitochondrial compartments (intermembrane space, inner membrane, or matrix) or with lysosomes or total cytosol. The antiserum was employed to characterize precursors of outer mitochondrial membrane proteins synthesized in vitro in a rabbit reticulocyte cell-free system. One product (a 68 kDa polypeptide designated OMM-68) bound efficiently to mitochondria in vitro but did not interact with either dog pancreas or rat liver microsomes, either co-translationally or post-translationally. OMM-68 was synthesized exclusively by the membrane-free class of polyribosomes. Attachment of precursor OMM-68 to mitochondria was not accompanied by processing of the polypeptide to a different size.     

Influence of diet and hypoxia on brain serotonin and catecholamines in rainbow trout (Salmo gairdneri)

1. In the brain of Salmo gairdneri, the content of dopamine (DA), norepinephrine (NE), serotonin (5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) depends upon the location in the brain (hypothalamus, telencephalon or mesencephalon). 2. The origin of feed protein (from animal or vegetal origin) influences the level of the various monoamines studied in different brain structures. 3. Hypoxia (60% oxygen saturation in water) causes modifications of 5-HT and catecholamine (DA, NE) contents in different brain structures, depending upon the diet.     

Biological tertiary treatment of urban wastewaters with chitosan-immobilizedPhormidium

Summary Chitosan:Phormidium aggregates (chitosan: algae=1:2, dry weight basis) were used as a biological tertiary treatment to remove the nitrogen (NH ₄ ⁺ , NO ₂ ^- , NO ₃ ^- ) and phosphorus (PO ₄ ^3- ) from a secondary effluent. In a batch system, 71 and 92% of P–PO ₄ ^3- were removed after 6 and 24 h, respectively. The orthophosphate removal rate was identical for all three concentrations of algae-chitosan tested (3.3, 4.6, 5.9 g d. wt.·l^-1), and was 90 g±2 g P–PO ₄ ^3- ·l^-1·h^-1, for a 90% removal. Under control conditions (chitosan flakes only added to the effluent) 73 and 78% of PO ₄ ^3- were removed after 6 and 24 h respectively. A 95% removal of inorganic nitrogen (NH ₄ ⁺ , NO ₂ ^- , NO ₃ ^- ) was attained after 4–6 h withPhormidium immobilized on chitosan flakes, as compared to 30% with chitosan flakes alone (5 g d. wt.·l^-1). The system gave a similar performance when operated semi-continuously over 5 days at a daily retention time of 1.0. In the presence of chitosan-immobilized algae, medium P–PO ₄ ^3- levels were reduced by 87.3%±6.4% after 24 h (61.1 g±7.0 g P·l^-1·h^-1). The reduction of inorganic nitrogen in the medium was 98% after 24 h (370 g±50 g N·l^-1·h^-1). In the presence of chitosan alone, some 60% orthophosphate removal was recorded, whereas no reduction of nitrogen was observed. Disappearance of orthophosphate was attributed to its co-precipitation with calcium released from the chitosan by abrasion. The presence of the algae protected the chitosan from abrasion andPhormidium directly assimilated the orthophosphate and inorganic nitrogen, thus reducing their levels in the effluent.     

True hermaphroditism in a child with a chimeric XX/XY chromosome, a double erythrocyte population and an unusual Lewis (a+ b+) phenotype

A 2 years-old Korean girl is seen because of ambiguous external genitalia. Surgical exploration shows the right gonad to be an ovary and the left one to be an ovotestis, thus demonstrating a true hermaphroditism. Cytogenetic studies of peripheral lymphocytes reveal a mixture of 46,XX and 46,XY cells, with a predominant XX cell line. The patient's red cells are composed of two distinct populations differing in three genetically independent blood group systems. The ratios of the two cell lines in various tissues, especially among the cells secreting Lewis antigens, appear to be very different and suggest several hypothesis to explain the highly unusual red cell Lewis phenotype Le (a+ b+). We conclude to a dispermic chimera, however the adopted status of this child prevents any identification of the maternal or paternal contributions. Because of the physical aspect it was decided to remove the ovotestis, to repair the external genitalia and to bring up this child as a female.     

抗—HBe阳性慢性乙肝临床研究

报告34例抗-HRe阳性慢性乙型肝炎(CHB),占同期住院抗-HBe阳性者的38％。这些患者于3—9年内肝炎再活动2—4次,累计80次。肝炎再活动时的临床表现及肝功损害与同期住院的HBeAg阳性CHB相似,但抗-HBe阳性CHB者肝硬化及肝癌发生比例显著高于HBeAg阳性者(P<0.05)。血清乙肝病毒标志(HBVM)检测发现,80次肝炎再活动中,38次(47.5％)有HBV活跃复制,提示HBV活跃复制是部分抗-HBe阳性CHB肝炎再活动的根本原因,另一部要考虑是其它肝炎病毒重叠感染的结果。