Amphiphilic cationic peptides mediate cell adhesion to plastic surfaces

Four amphiphilic peptides, each with net charges of +2 or more at neutrality and molecular weights under 4 kilodaltons, were found to mediate the adhesion of normal rat kidney fibroblasts to polystyrene surfaces. Two of these peptides, a model for calcitonin (peptide 1, MCT) and melittin (peptide 2, MEL), form amphiphilic alpha-helical structures at aqueous/nonpolar interfaces. The other two, a luteinizing hormone-releasing hormone model (peptide 3, LHM) and a platelet factor model (peptide 4, MPF) form beta-strand structures in amphiphilic environments. Although it contains only 10 residues, LHM mediated adhesion to surfaces coated with solutions containing as little as 10 pmoles/ml of peptide. All four of these peptides were capable of forming monolayers at air-buffer interfaces with collapse pressures greater than 20 dynes/cm. None of these four peptides contains the tetrapeptide sequence Arg-Gly-Asp-Ser, which has been associated with fibronectin-mediated cell adhesion. Ten polypeptides that also lacked the sequence Arg-Gly-Asp-Ser but were nonamphiphilic and/or had net charges less than +2 at neutrality were all incapable of mediating cell adhesion (Pierschbacher and Ruoslahti, 1984). The morphologies of NRK cells spread on polystyrene coated with peptide LHM resemble the morphologies on fibronectin-coated surfaces, whereas cells spread on surfaces coated with MCT or MEL exhibit strikingly different morphologies. The adhesiveness of MCT, MEL, LHM, and MPF implies that many amphiphilic cationic peptides could prove useful as well defined adhesive substrata for cell culture and for studies of the mechanism of cell adhesion.     

Pollarding and a possible explanation of the neolithic elmfall

Sections of two pollarded parkelms (Ulmus glabra) from Damsgaard, Bergen, west Norway have been studied. Changes in annual ring-width are attributed partly to management, namely pollarding, and partly to pathogenic attacks, probably by Ceratocystis ulmi. The oldest attack on the trees dates back to 1826: so far the oldest known record of Dutch elm disease in Norway. Pollarding may be an important factor in attacks by the pathogen within parkelms. A possible relationship between pollarding, the pathogen and the Neolithic elmfall is suggested.     

The catalytic mechanism of bovine intestinal 5'-nucleotide phosphodiesterase. pH and inhibition studies

Extensive kinetic studies of bovine intestinal 5'-nucleotide phosphodiesterase as a function of pH have confirmed and amplified the catalytic mechanism previously proposed on the basis of isolation of a covalent phosphorylated intermediate (Landt, M., and Butler, L.G. (1978) Biochemistry 17, 4130-4135). An enzyme-ionizing group with apparent pKa = 6.85 controls the rate-determining step. Electrostatic interactions between anionic substrate and two or more ionic groups on the enzyme have a major role in substrate binding. Binding of strongly inhibitory 5'-AMP is controlled by an ionizing group, probably on the enzyme, with pKa less than or equal to 5.9. At pH 6.0, imidazole is a classic uncompetitive inhibitor, in agreement with independent evidence that it stabilizes the covalent intermediate form of the enzyme. KI values for phosphonate analogs, which are competitive inhibitors, indicate that phosphodiesterase binds its products and product analogs more strongly than it binds substrate analogs. Some of the results presented here can be interpreted as indicating that 5'-nucleotide phosphodiesterase is the evolutionary precursor of alkaline phosphatase, with which it has many structural and catalytic properties in common, and which is found in relatively large amounts in the same tissue.     

在我国腹泻患儿中发现诺瓦克样病霉感染

诺瓦克病毒是引起无菌性急性胃肠炎爆发的重要病原。１９９０年１０月至１９９１年１月从河南省急性腹泻门诊患儿便样中发现了诺瓦克样病毒。经电镜观察,病毒直径约为２８ｎｍ,病毒壳似由有结构的亚单位组成,进一步用诺瓦克病毒特异的寡核苷酸引物做逆转录一聚合酶链式反应（ＲＴ－ＰＣＲ）,检定为阳性。由ＰＣＲ产物测得的核苷酸序列与诺瓦克病毒原型株相应序列比较,同源性为７２％。以上结果证实,在我国腹泻病人中有诺瓦克样病毒感染。这对我国病毒性胃肠炎流行的防治研究具有重要意义。     

A new approach for estimating the crosslink density of covalently crosslinked ionic polysaccharide gels

For an ideal polysaccharide gel with a known total polymer chain contour length, crosslinks all of the same functionality and elastic chains all with the same contour length and stiffness, the gel crosslink density can readily be determined from measurements of the maximum volume of the swollen gel (Moe et al., (1991) Food Hydrocolloids, 5, (1/2), 119–123. In the case of randomly crosslinked polysaccharide gels, where the chain contour length between two adjacent crosslinks may vary greatly, it is often much more difficult to determine the crosslink density. This paper reports on an attempt to extend the use of maximum gel volume measurements to estimate crosslink density for the latter type of gel. This is done by calculating the maximum swelling volume for polymer networks with four-functional crosslinks, known elastic chain mean contour length and standard deviation. The numerical analysis involves the calculation of the equilibrium force at each crosslink as the network expands. This allows a detailed study of how the distribution of individual polymer chain contour lengths affects the maximum swelling volume. The computer simulation results are compared with the results from experimental measurements of the maximum volume of swollen covalently crosslinked sodium alginate gels.     

Molecular cut-off values of dialysis membranes for alginate and kappa-carrageenan oligosaccharides

The effective molecular weight cut-off values of dialysis membranes for carrageenan and alginate oligosaccharides were evaluated by gel permeation chromatography and nuclear magnetic resonance spectroscopy. For the different membranes tested, i.e. Medicell, Spectra Por 1000D and 3500D, the porous sizes are analogous to tri- and tetrasaccharides. A simple dialysis can be used to recover the majority of the oligosaccharides produced by a carrageenase or an alginate lyase digestion.     

Biotin carboxylases in mitochondria and the cytosol from skeletal and cardiac muscle as detected by avidin binding

Biotin carboxylases in mammalian cells are regulatory enzymes in lipogenesis and gluconeogenesis. In this study, endogenous biotin in skeletal and cardiac muscle was detected using avidin conjugated with alkaline phosphatase and applied in high concentrations to muscle sections. The avidin binding was subsequently visualized by histochemical demonstration of the alkaline phosphatase activity. All cardiac muscle cells showed high affinity for avidin with only the nuclei and the intercalated discs remaining unstained. In skeletal muscle a diffuse reaction could be detected in the sarcoplasm of the muscle fibres. A granular reaction was noted in the same fibres that showed activity for succinic dehydrogenase. The specificity of the coloured reaction product in the muscle sections was investigated and is suggested to be caused by avidin binding to biotin moieties in mitochondria and the cytosol. Mitochondrial and cytosolic preparations of skeletal muscle were electrophoresed in sodium dodecyl sulphate gels. After blotting and incubation with conjugated avidin, two bands with molecular weights of 75 kDa and 130 kDa respectively were evident in the mitochondrial preparation. It is suggested that the 75-kDa band represents comigration of the biotin-containing subunits of propionyl-CoA carboxylase and methylcrotonyl-CoA carboxylase. The 130-kDa band may represent the biotin-containing pyruvate carboxylase. In the cytosolic preparation a 270-kDa band was stained in blots that had been incubated with conjugated avidin; this band is suggested to represent acetyl-CoA carboxylase. A 190-kDa cytosolic band might be a cleavage product of acetyl-CoA carboxylase. We propose that using alkaline phosphatase-conjugated avidin it is possible to detect the mitochondrial and cytosolic biotin-dependent carboxylases in striated muscle.     

Circular dichroism of the parallel β helical proteins pectate lyase C and E

The pectate lyases, PelC and PelE, have an unusual folding motif, known as a parallel β-helix, in which the polypeptide chain is coiled into a larger helix composed of three parallel β-sheets connected by loops having variable lengths and conformations. Since the regular secondary structure consists almost entirely of parallel β-sheets these proteins provide a unique opportunity to study the effect of parallel β-helical structure on circular dichroism (CD). We report here the CD spectra of PelC and PelE in the presence and absence of Ca²⁺, derive the parallel β-helical components of the spectra, and compare these results with previous CD studies of parallel β-sheet structure. The shape and intensity of the parallel β-sheet spectrum is distinctive and may be useful in identifying other proteins that contain the parallel β-helical folding motif. © 1995 Wiley-Liss, Inc.