Purification of atrial natriuretic peptide receptor from bovine lung. Evidence for a disulfide-linked subunit structure

The receptor for atrial natriuretic peptide (ANP) was purified to apparent homogeneity from bovine lung by a combination of detergent extraction, ammonium sulfate precipitation, and affinity chromatography on ANP-Affi-Gel 10. The Mr of the purified receptor is about 140,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. After reduction, the protein migrated as a single band with an Mr near 70,000. NH2-terminal sequence analysis of the purified material revealed only one sequence, indicating that the ANP receptor is composed of two probably identical subunits held together by disulfide bond(s), although it remains possible that one of the subunits is blocked at the NH2 terminus. Antibody was produced to the nonreduced Mr = 140,000 species and shown to interact with detergent-solubilized forms of the lung and kidney ANP receptor.     

Fibrinolysis by urokinase endowed with magnetic property

The activated magnetic modifier was synthesized from magnetite, alpha, omega-dicarboxymethylpoly(oxyethylene) and N-hydroxysuccinimide (Biochem. Biophys. Res. Commun., 145, 908-914, 1987). Urokinase was directly coupled with the activated magnetic modifier to obtain magnetic urokinase. The magnetic urokinase dispersed in saline and exerted high fibrinolytic activity (13.8 X 10(4) IU/mg protein), and was readily recovered from saline by magnetic force of 250 Oe. By applying magnetic force, the urokinase was attracted at our will and local fibrinolysis was achieved on fibrin gel in a petri dish.     

Secondary release of thromboxane A2 in aerosol leukotriene C4-induced bronchoconstriction in guinea pigs

Effect of a thromboxane synthetase inhibitor (OKY-046) on bronchoconstriction induced by aerosol leukotriene C4 and histamine was studied in anesthetized, artificially ventilated guinea pigs in order to examine whether secondary release of thromboxane A2 is produced by aerosol leukotriene C4 or not. 0.01–1.0μg/ml of leukotriene C4 and 12.5–400μg/ml of histamine inhaled from ultrasonic nebulizer developed for small animals caused dose-dependent increase of pressure at airway opening (Pao) which is considered to be an index representing bronchial response. Pretreatment of the animals with intravenous OKY-046 (100mg/kg) significantly reduced the airway responses produced by inhalation of 0.1, 0.33 and 1.0μg/ml of leukotriene C4, while the pretreatment did not affect the histamine dose-response curve. Based on these findings and previous reports (6, 7), it is suggested that aerosol leukotriene C4 activates arachidonate cyclooxygenase pathway including thromboxane A2 synthesis and the released cyclooxygenase products have bronchodilating effect as a whole     

Structure of the Bombyx mori fibroin light-chain-encoding gene: upstream sequence elements common to the light and heavy chain.

Two overlapping genomic clones containing the fibroin light-chain (Fib-L)-encoding gene (Fib-L) were obtained from the cosmid library of the silkworm, Bombyx mori J-139, by hybridization with the Fib-L cDNA clone. Sequencing of the 14.6-kb region revealed that Fib-L was 13472 bp long containing seven exons, and that the gene contained a large first intron which occupied about 60% of the gene. Comparison of restriction patterns of the J-139 Fib-L with those of eight other B. mori breeds producing normal-level fibroin demonstrated that considerable restriction-fragment length polymorphisms were present in regions containing the first intron and the 3′-flanking sequence. However, sizes of the Fib-L mRNA and the Fib-L polypeptide were very similar among the nine breeds tested, suggesting that the exon sequences and the splice signals were all well conserved. 5′-Flanking regions of Fib-L and the fibroin heavy-chain (Fib-H)-encoding gene (Fib-H) compared in this study contained three 18-30-bp sequences of high similarity and many 8-10-bp common elements, six of which coincided with the binding sites of homeodomain proteins. Gel retardation assays with the nuclear extracts of the posterior and middle silk glands suggested that protein factors present in the posterior silk-gland nuclei could bind to a set of those common upstream elements.     

A leptomycin B resistance gene of Schizosaccharomyces pombe encodes a protein similar to the mammalian P-glycoproteins

Screening for leptomycin B (LMB)-resistant transformants in a gene library constructed in Schizosaccharomyces pombe with the chromosomal DNA of an LMB-resistant mutant of S. pombe and with multicopy plasmid pDB248' as the vector led to the isolation of a gene, named pmd1+, encoding a 1362-amino-acid protein. This protein showed great similarity in amino acid sequence to the mammalian P-glycoprotein encoded by the multidrug resistance gene, mdr, and the Saccharomyces cerevisiae a-factor transporter encoded by STE6. In addition, computer analyses predicted that the protein encoded by pmd1+ formed an intramolecular duplicated structure and each of the halves contained six transmembrane regions as well as two ATP-binding domains, as observed with the P-glycoproteins and the STE6 product. Consistent with this was that S. pombe cells containing the pmd1+ gene on a multicopy plasmid showed resistance not only to LMB but also to several cytotoxic agents. The pmd1 null mutants derived by gene disruption were viable and hypersensitive to these agents. All these data suggest that the pmd1+ gene encodes a protein that is a structural and functional counterpart of mammalian mdr proteins.     

Endotoxin and cytokines in patients with gastrointestinal tract perforation

Plasma levels of endotoxin and various cytokines were assessed in 70 patients with gastrointestinal tract perforation. Sepsis developed in 29 of them, and eight of these (27.6%) had on admission endotoxin levels higher than 9.8 pg ml(-1). The clinical outcome correlated with the level of tumour necrosis factor alpha (TNFalpha), rather than with the endotoxin level. The high interleukin 6 (IL-6) level was shown in septic patients and no correlation was observed between the IL-6 level and the clinical outcome. Plasma TNFalpha levels tended to change independently from endotoxin levels, suggesting that TNFalpha may have been locally produced in inflammatory lesions.