The interaction of collagen with alpha1-acid glycoprotein.

The influence of alpha1-acid glycoprotein on the formation of fibrous long spacing fibers of collagen has been investigated. It was observed that addition of the glycoprotein to dialyzed collagen solutions caused a significant decrease in the intensity of the circular dichroic spectrum of collagen. This phenomenon, which displays an optimum with respect to glycoprotein, is consistent with previous observations of fibrous long spacing fiber formation. Changes in viscosity of collagen initially dissolved in acetic acid were monitored during dialysis. It was found that a significant increase in viscosity must occur during dialysis of collagen before fibrous long spacing formation could take place. This increase in viscosity can be related directly to removal of acetic acid from the collagen solution. Removal of all sialyl residues from the alpha1-acid glycoprotein with neuraminidase prevents fibrous long spacing formation while removal of up to 35% of the sialyl residues has no effect on the interaction of glycoprotein with collagen. Amino acid composition and radioactivity studies suggest that 45-55% of the insoluble fibrous long spacing fibers is glycoprotein. In contrast to native collagen fibers, reduced fibrous long spacing fibers do not contain histidinohydroxymerodesmosine or hydroxylysinonorleucine. Instead, they contain significant quantities of allysine aldol and epsilon-hydroxynorleucine.     

Zur Kenntnis der Farbstoffresorption durch die Darmschleimhaut

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Changes in the cyclic nucleotide level and the inhibition of human thrombocyte aggregation in 3-hydroxypyridine exposure

The effect of six 3-oxypiridine derivatives (at a concentration of 10(-3) and 5 X 10(-3) M) on cyclic nucleotide level in human platelets and platelet aggregation was studied. Five 3-oxypiridine derivatives were shown to depress platelet aggregation, four of them causing the increase in cAMP platelet level. The correlation between antiaggregation activity of 3-oxypiridine derivatives and their ability to rise cyclic nucleotide level in human platelets is discussed.     

Amphipathic beta structure of a leucine-rich repeat peptide.

Long tandem arrays of a characteristic leucine-rich repeat motif on the order of 24 amino acids in length have been found in the primary structure of an increasing number of proteins. The most striking feature of these repeats is an amphipathic sequence, with leucine as the predominant hydrophobic residue. Based on this amphipathic sequence and the function of the proteins in which they have been found, the repeats have been proposed to be involved in protein-protein and protein-lipid interactions. As a step toward elucidating the structure and biochemical properties of the leucine-rich repeat motif, we have studied a synthetic leucine-rich repeat peptide (LRP32) representing one of the repeats found in Drosophila chaoptin. We have shown that: (i) LRP32 is soluble in aqueous solution but will bind quantitatively to phospholipid vesicles; (ii) LRP32 has a partial beta structure in aqueous solution and is predominantly a beta structure in the presence of phospholipid; (iii) LRP32 integrates into lipid bilayers to form 60-A intramembrane particles as seen using freeze-fracture electron microscopy (these putative oligomeric structures appear to contain a central aqueous core as indicated by their ability to generate conductances in planar lipid bilayers); and (iv) LRP32-lipid complexes generate 2H NMR spectra characteristic of integral membrane proteins. This study is consistent with LRP32 forming an amphipathic beta sheet. We propose that protein segments containing tandem arrays of leucine-rich repeats also may form amphipathic beta sheets.     

Mechanisms of the effects of Ca2+ ions on lipid peroxidation

Mechanisms underlying Ca2+ effects on lipid peroxidation (LPO) induced in liposomes (from egg yolk lecithin) and UFsomes (from linolenic acid, methyl linolenate) with the aid of O2- -system (Fe2+ + ascorbate) were studied. It was shown that stimulation of lipid peroxidation by low Ca2+ concentrations (10(-6)-10(-5) M) was due to its ability to release Fe2+-ions bound to negatively charged (phosphate, carboxylic) lipid groups (of licethin, linolenic acid), thus increasing the concentration of catalytically active Fe2+. The inhibitory effect of high Ca2+ concentrations was caused by its interaction with superoxide anion-radicals and was not observed in LPO-systems, independent of O2- generation (e. g. Fe2+ + cumol hydroperoxide).     

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Activation and mechanism of Clostridium septicum alpha toxin

Clostridium septicum produces a single lethal factor, alpha toxin (AT), which is a cytolytic protein with a molecular mass of approximately 48kDa. The 48kDa toxin was found to be an inactive protoxin (AT^pro) which could be activated via a carboxy-terminal cleavage with trypsin. The cleavage site was located approximately 4kDa from the carboxy-terminus. Proteolytically activated AT^pro had a specific activity of approximately 1.5 × 10⁶ haemolytic units mg^-1. The trypsin-activated toxin (AT^act) was haemolytic, stimulated a prelytic release of potassium ions from erythrocytes which was followed by haemoglobin release, induced channel formation in planar membranes and aggregated into a complex of M_r >210000 on erythrocyte membranes. AT^pro did not exhibit these properties. AT^act formed pores with a diameter of at least 1.3-1.6 nm. We suggest that pore formation on target cell membranes is responsible for the cytolytic activity of alpha toxin.     

Long-chain Acyl-CoA Dehydrogenase Deficiency as a Cause of Pulmonary Surfactant Dysfunction

Long-chain acyl-CoA dehydrogenase (LCAD) is a mitochondrial fatty acid oxidation enzyme whose expression in humans is low or absent in organs known to utilize fatty acids for energy such as heart, muscle, and liver. This study demonstrates localization of LCAD to human alveolar type II pneumocytes, which synthesize and secrete pulmonary surfactant. The physiological role of LCAD and the fatty acid oxidation pathway in lung was subsequently studied using LCAD knock-out mice. Lung fatty acid oxidation was reduced in LCAD^−/− mice. LCAD^−/− mice demonstrated reduced pulmonary compliance, but histological examination of lung tissue revealed no obvious signs of inflammation or pathology. The changes in lung mechanics were found to be due to pulmonary surfactant dysfunction. Large aggregate surfactant isolated from LCAD^−/− mouse lavage fluid had significantly reduced phospholipid content as well as alterations in the acyl chain composition of phosphatidylcholine and phosphatidylglycerol. LCAD^−/− surfactant demonstrated functional abnormalities when subjected to dynamic compression-expansion cycling on a constrained drop surfactometer. Serum albumin, which has been shown to degrade and inactivate pulmonary surfactant, was significantly increased in LCAD^−/− lavage fluid, suggesting increased epithelial permeability. Finally, we identified two cases of sudden unexplained infant death where no lung LCAD antigen was detectable. Both infants were homozygous for an amino acid changing polymorphism (K333Q). These findings for the first time identify the fatty acid oxidation pathway and LCAD in particular as factors contributing to the pathophysiology of pulmonary disease.