Sequence of a full-length cDNA for rat lung beta-galactoside-binding protein: primary and secondary structure of the lectin

A full-length cDNA for rat lung beta-galactoside lectin (subunit Mr approximately 14,000, lectin 14K) was cloned and the nucleotide sequence determined. The deduced amino acid sequence agrees with the amino acid composition and direct amino acid sequence analysis of purified rat lung lectin peptides. We found that the amino-terminal alanine is blocked with an acetyl group. Comparison of the amino acid sequence with other proteins shows a high degree of homology only with other vertebrate lectin sequences, supporting the suggestion that these lectins may constitute a unique class of vertebrate proteins. The amino acid composition and sequence of lectin peptides, the sequence of lectin cDNA, and isoelectric focusing of purified lectin indicate that rat lung lectin 14K is composed predominantly of a single protein. In addition, rat uterus lectin 14K was found to be the same protein as that present in lung. We characterized the secondary and tertiary structure of rat lung lectin 14K by circular dichroism, by analytical ultracentrifugation, and by computer analysis of its primary structure. Results of these experiments suggest that lectin 14K is primarily a hydrophilic protein with an asymmetric, elongated structure consisting of approximately equal amounts of alpha helix, beta sheet, beta turn, and random coil. We found that Cys-2 and Cys-130 react most rapidly with iodoacetamide; one or both of these residues may be primarily responsible for the thiol requirement of lectin activity.     

Growth hormone releasing factor-like immunoreactivity in human milk

The presence of immunoreactive growth hormone-releasing factor (GRF) in human milk has been demonstrated. By using sequential high performance liquid chromatography, it has been shown that most of the immunoreactivity co-elutes with the synthetic, hypothalamic-like, GRF (1-40). The concentrations of GRF detected (between 152 and 432 pg GRF/ml milk) exceed several fold its values in plasma.     

Downregulation of cell-to-cell communication by the viralsrc gene is blocked by TMB-8 and recovery of communication is blocked by vanadate

Summary The viralsrc gene downregulates junctional communication, closing cell-to-cell membrane channels presumably by way of the phosphoinositide signal route. We show that TMB-8 [8-N, N-(diethylamino) octyl-3,4,5-trimethoxybenzoate] counteracts this downregulation in cells transformed by temperature-sensitive mutant Rous sarcoma virus: TMB-8 (36–72 m) raises junctional permeability when applied during activity ofsrc protein kinase, i.e., at steady permissive temperature; and TMB-8 inhibits the fall of junctional permeability, when the activity ofsrc protein kinase gets turned on. TMB-8 also (reversibly) inhibits the growth of the cells at permissive temperature and reverses the morphological changes associated with transformation. The morphological reversal lags several hours behind the junctional-permeability reversal. Communication recovers within a few minutes when the activity of thesrc protein kinase is turned off (in absence of TMB-8). Sodium orthovanadate (20 m) prevents this recovery, but it has no major effect on junctional permeability on its own. We discuss possible modes of action of these agents on critical stages of the signal route, related to intracellular Ca²⁺ and protein kinase C.     

Seminal prostaglandins in men with subnormal sperm motility and therapeutic treatment

The contents of prostaglandins in seminal plasma from a total of 73 men were evaluated. The subjects were grouped as follows: normospermic men, patients with impaired motility, patients with small untreated varicocelle and patients with impaired motility and Kallikrein therapy. Sperm density, morphology and motility were examined. High performance reversed phase liquid chromatography (HPLC) in combination with specific radioimmunoassays were used for the determination of PGE₂, PGI₂ and PGF_2α. There was a significant difference (p < 0, 025; F-test) between the PGI₂ concentrations in patients with impaired motility (5,6 ± 1,4 pg/mg protein) and normal men (8,8 ± 3,7 pg/mg protein). PGE₂ and PGF_2α were significantly different in patients with varicocele (p < 0,025, F-test). Wide ranges of prostaglandins occured in the Kallikrein-group with no significant differences. We conclude that: a) PGI₁ is an additional prostaglandin compound in seminal plasma. b)its measurement may not be useful as diagnostic parameter in subfertile men and c) Kallikrein has no influence on the prostaglandin content in seminal plasma and other seminal parameters as motility, motility index and sperm counts.     

Plathelminth abundance in North Sea salt marshes: environmental instability causes high diversity

Although supralittoral salt marshes are habitats of high environmental instability, the meiofauna is rich in species and abundance is high. The community structure of free-living Plathelminthes (Turbellaria) in these salt marshes is described. On an average, 104 individuals are found below an area of 10 cm². The average species density in ungrazed salt marshes is 11.3 below 10 cm² and 45.2 below 100 cm², indicating strong small-scale heterogenity. The faunal similarity between sediment and the corresponding above-ground vegetation is higher than between adjacent sample sites. Species prefer distinct ranges of salinity. In the lower part of the supralittoral salt marshes, the annual fluctuations of salinity are strongest and highly unpredictable. This region is richest in plathelminth species and abundance; diversity is highest, and the faunal composition of parallel samples is quite similar. In the upper part of the supralittoral salt marshes, the annual variability of salinity is lower, plathelminths are poor in species diversity and abundance. Parallel samples often have no species in common. Thus, those salt marsh regions with the most unstable environment are inhabited by the most diverse species assemblage. Compared to other littoral zones of the North Sea, however, plathelminth diversity in salt marshes is low. The observed plathelminth diversity pattern can apparently be explained by the dynamic equilibrium model (Huston, 1979).     

Effect of long-term anemia and retransfusion on central circulation during exercise

The bulk modulus and the shear modulus describe the capacity of material to resist a change in volume and a change of shape, respectively. The values of these elastic coefficients for air-filled lung parenchyma suggest that there is a qualitative difference between the mechanisms by which the parenchyma resists expansion and shear deformation; the bulk modulus changes roughly exponentially with the transpulmonary pressure, whereas the shear modulus is nearly a constant fraction of the transpulmonary pressure for a wide range of volumes. The bulk modulus is approximately 6.5 times as large as the shear modulus. In recent microstructural modeling of lung parenchyma, these mechanisms have been pictured as being similar to the mechanisms by which an open cell liquid foam resists deformations. In this paper, we report values for the bulk moduli and the shear moduli of normal air-filled rabbit lungs and of air-filled lungs in which alveolar surface tension is maintained constant at 16 dyn/cm. Elevating surface tension above normal physiological values causes the bulk modulus to decrease and the shear modulus to increase. Furthermore, the bulk modulus is found to be sensitive to a dependence of surface tension on surface area, but the shear modulus is not. These results agree qualitatively with the predictions of the model, but there are quantitative differences between the data and the model.     

Molecular characterization of the human erythrocyte anion transport protein in octyl glucoside

Band 3 protein, the anion transport protein of the human erythrocyte membrane, was purified in the presence of the nonionic detergent octyl glucoside. A molecular characterization was carried out to investigate whether the native structure of the protein was retained in the presence of this detergent. Band 3 bound octyl glucoside below the critical micelle concentration (cmc) of the detergent, approaching saturation above the cmc. At 40 mM octyl glucoside, close to saturating concentrations, 0.64 g of octyl glucoside is bound per gram of band 3 protein, corresponding to 208 molecules of detergent bound per monomer of band 3. Sedimentation velocity and gel filtration studies, performed at 40 mM octyl glucoside, indicated that the band 3-octyl glucoside complex had an average molecular weight of 1.98 X 10(6), which corresponds to a dodecamer. Sedimentation equilibrium experiments confirmed that band 3 in octyl glucoside exists in a heterogeneous and high oligomeric state. This high oligomeric state did not change dramatically over octyl glucoside concentrations ranging from 6 to 60 mM. The circular dichroism spectrum of band 3 changed only slightly over this range of octyl glucoside concentrations. The alpha-helical and beta-sheet contents of band 3 in 2 mM octyl glucoside were calculated to be 40% and 27%, respectively, indicating that no gross alteration in the secondary structure of the protein had occurred in octyl glucoside. The ability of band 3 to bind 4-benzamido-4'-aminostilbene-2,2'-disulfonate (BADS), a potent inhibitor (Ki = 1 microM) of anion transport, was measured to assess the integrity of the inhibitor binding site of the protein in octyl glucoside.(ABSTRACT TRUNCATED AT 250 WORDS)     

Purification of a phospholipase B inhibitor from Saccharomyces cerevisiae

The phospholipase B activity of plasma membrane vesicles from Saccharomyces cerevisiae is inhibited by the 100 000 X g supernatant of mechanically disrupted yeast cells. A 1850-fold purification of the inhibitor activity was achieved by gel filtration, ion exchange chromatography with DEAE-cellulose and hydrophobic interaction chromatography with Octyl-Sepharose. SDS-polyacrylamide gel electrophoresis of the purified inhibitor revealed two main bands with an apparent Mr of 60 000 and 26 500. The phospholipase B activity was strongly reduced but not completely blocked by this preparation, while the lysophospholipase and transacylase reactions, which are catalyzed by the same membrane-bound enzymes (Witt, W. et al. (1984) Biochim. Biophys. Acta 795, 108-116), were not affected.     

Amino acid sequence of an invertebrate FBP aldolase (from Drosophila melanogaster)

The complete amino acid sequence of FBP aldolase from Drosophila melanogaster has been determined. The enzyme contains four identical subunits of 360 amino acid residues. The primary structure of the monomer was established using automated Edman degradation on fragments prepared by CNBr-cleavage, by partial acid cleavage at the unique Asp-Pro bond and by oxidative cleavage at the three tryptophan residues. Manual Edman-Chang degradation was used on smaller peptides obtained by digestion with Staphylococcus aureus V8 protease, trypsin or chymotrypsin. The primary structure of Drosophila aldolase exhibits very extensive homology with the sequence of rabbit muscle aldolase (71% identity), thus explaining the early observation that Drosophila and mammalian aldolases form active interspecies hybrid quaternary structures (Brenner-Holzach, O. and Leuthardt, F., Eur. J. Biochem. (1972) 31, 423-426).     

Translation and functional expression of cell-cell channel mRNA inXenopus oocytes

Summary mRNA from estrogen-stimulated rat myometrium, a tissue known to upregulate cell-cell channels in response to this hormone, was microinjected intoXenopus laevis oocytes. The oocytes had been freed from covering layers of follicle cells and vitelline to allow direct cell membrane interactions when paired. About 4 hours after the mRNA injection, paired oocytes become electrically coupled. This coupling was due to the presence of typical cell-cell channels characterized by size-limited intercellular tracer flux, the presence of gap junctions at the oocyte-oocyte interface, and the reversible uncoupling that occurred in the presence of carbon dioxide. The induction of new cell-cell channels in the oocyte membrane was observed against a zero background or a low level of endogenous coupling, depending on the maturation stage of the oocytes. The time course of development of cell-cell coupling after the microinjection of mRNA was determined. The mRNA capable of inducing cell-cell coupling was confined to an intermediate size class when fractionated on a sucrose gradient.