Solution conformation of Lewis a-derived selectin ligands is unaffected by anionic substituents at the 3'- and 6'- positions

The selectins are a family of proteins that mediate leukocytetethering and rolling along the vascular endothelium. E-, P-,and L-selectin recognize various derivatives of the Lewis^a andLewis^x trisaccharides. The distribution of negative chargeson the Lewis^a and Lewis^x oligosaccharides appears to be an importantfactor in their binding by the selectins. Previous work exploringthis electrostatic dependence found that a series of syntheticanionic trisaccharides, 3'-sulfo, 3'-phospho, 6'-sulfo, and3',6'-disulfo Lewis^a. (Glc), exhibited differing selectin inhibitoryefficacies. To explore the possibility that these differencesarise from conformational differences between the sugars, thesolution structures of these trisaccharides were determinedusing NMR and molecular dynamics simulations. Interproton distancesand interglycosidic torsion angles were determined at 37°Cusing NOESY buildup curves and 1D LRJ experiments, respectively.Data from both experiments agreed well with predictions madefrom 2000 picosecond unrestrained molecular dynamics simulations.We found that 3'-sulfation did not alter the core Lewis^a conformation,a finding that reaffirms the results of previous study. In addition,we found that sulfation at the 6' position also leaves the trisaccharideconformation unperturbed. This is significant because the proximityof the 6'-sulfate group to the fucose ring might have alteredthe canonical Lewis^a structure. The disulfate exhibited greaterflexibility than the other derivatives in dynamics simulations,but not so much as to affect NOE and heteronuclear couplingconstant measurements. Taken together, our findings supportthe use of Lewis^a as a template onto which charged groups maybe added without significantly altering the trisaccharide'sstructure. oligosaccharides molecular dynamics simulations NMR sulfated Lewis^a phosphorylated Lewis^a     

Reduction of vanadate to vanadyl by a strain of Saccharomyces cerevisiae

Three strains of Saccharomyces cerevisiae, SC-1, DBVPG 6173 and DBVPG 6037, were studied for vanadate resistance in complex Sabouraud medium since they did not thrive in different minimal media (yeast nitrogen base with and without amino acids). The strain SC-1 was resistant up to 16 mm of vanadate, whereas the strains DBVPG 6173 and DBVPG 6037 were inhibited by 8 mm and 4 mm vanadate, respectively. The vanadate resistance in strain SC-1 was constitutive and due to the reduction of this oxyanion to vanadyl, which was detected by EPR spectroscopy and visible spectroscopy. The transformation of vanadate to vanadyl took place during the exponential growth phase; 10 mm of vanadate was reduced to vanadyl outside the cells since the oxyanion was not detected in the cell biomass and only a negligible concentration of vanadyl (25 nmoles mg cells dry weight) was found in the biomass. The other two vanadate-sensitive yeast strains only accumulated vanadate and did not reduce the oxyanion to vanadyl.     

Two isoaccepting seryl tRNAs coded by separate mitochondrial genes in yeast

Summary In S. cerevisiae four isoacceptor mitochondrial tRNAs for serine have been separated by reversed phase chromatography. At least two of these species are products of different genes. In this work the deletion mapping technique has been used to locate two genes for tRNA^ser. The gene for tRNA^ser previously localized in the oli I region of the mitochondrial genome has been found to code for tRNA _ser ² , and another gene coding for tRNA _ser ¹ has been detected in the region where most of other tRNA genes are found. Results of fine mapping experiments allowed to localize this gene in the proximity of the gene for tRNA^arg.     

Actin organization and fibrin-clot retractile activity of cultured mouse fibroblasts

Summary It is known that human and animal fibroblasts are able to induce the retraction of a fibrin clot. In the present study the correlation between (i) fibrinclot retractile (FCR) activity, (ii) the number of actin stress-lines in mouse fibroblasts during growth in culture, and (iii) the sensitivity of actin stress-lines to a powerful actin-depolymerizing factor (ADF), present in plasma and serum of humans and laboratory animals was investigated. Fibroblasts at early passages (2–4) were tested for these parameters at various intervals after seeding (24, 96, and 168 h). The number of actin stress-lines was progressively higher, while the sensitivity to ADF action was progressively lower in cells cultured from 24 to 168 h; the FCR capacity was significantly decreased at 168 h. These data suggest that cells containing weakly polymerized and/or stabilized actin are more active than those containing highly polymerized and/or stabilized actin in triggering fibroblast contraction.     

Complex and simple sequences in human repeated DNAs

Highly repeated human DNA sequences were isolated by isopycnic centrifugation, or were eluted from gels after restriction enzyme cleavage. High molecular weight DNA peaks separable from the bulk of the DNA in a variety of gradients were shown to consist of very simple sequences characteristic of simple satellite DNAs; DNA fingerprint studies indicated each of these peaks could consist of tandem repeats of a specific oligonucleotide sequence as low as 10 base pairs (bp) long. All the gradient peaks could be assigned to one of two sequence groups and several different buoyant density peaks revealed the same sequence. — Restriction fragment multimers did not share common sequences with the satellite DNAs as judged by hybridization data. They could not be separated by isopycnic centrifugation. Furthermore these highly repeated DNAs were more complex in sequence and more variable than the satellites. Even the smallest (50 bp) fragments by depurination and other direct sequencing methods were shown to be more complex than the high molecular weight satellite peaks. — The idea that subsets of repeated DNAs may be defined by sequence complexity, possibly with discrete or separable functions, is proposed.     

Annulate lamellae in hindgut epithelial cells ofIsopoda

Summary Hindgut epithelial cells of the aquatic isopod,Asellus communis, and the terrestrial isopod,Armadillidium vulgare, possess annulate lamellae. The organelle is present in non-dividing cells of intermolt adult organisms. The lamellae exhibit dense pore areas traversed by diaphragms, and the lamellae are associated with elements of endoplasmic reticulum.This research was supported in part by a grant to E. R. Witkus from the Irene Heinz and John La Porte Given Foundation.     

Dynamics of the human heart rate during changes of posture under various climatic conditions]

The time course of the heart rate (15-sec-periods) during active orthostasis (10 min) was investigated according to types found by Drischel et al. under different climatic conditions in 51 female and 50 male subjects. Constant types were observed in each subject in 3 repeated tests on different days at 20 degrees C and 55% R.H., the types remaining constant even at 25 and 90% R.H. The increase in environmental temperature from 15 to 30 and 45 degrees C showed a significant alteration from type A via different transitional types to type B. At 45 degrees C, an additional type was observed called superhigh type B, functionally being a precollapse type. The variation in typology includes the dynamics of the initial process of the active orthostasis as well as the deviation between the steady-state of the heart rate in erect and in supine position. The physiological mechanisms eliciting the changes in the dynamic behaviour and the correlations with the effective temperature are discussed.     

Proteomic analysis of microvesicles from plasma of healthy donors reveals high individual variability

Healthy blood plasma is required for several therapeutic procedures. To maximize successful therapeutic outcomes it is critical to control the quality of blood plasma. Clearly initiatives to improve the safety of blood transfusions will have a high economical and social impact. A detailed knowledge of the composition of healthy blood plasma is essential to facilitate such improvements. Apart from free proteins, lipids and metabolites, blood plasma also contains cell-derived microvesicles, including exosomes and microparticles from several different cellular origins. In this study, we have purified microvesicles smaller than 220nm from plasma of healthy donors and performed proteomic, ultra-structural, biochemical and functional analyses. We have detected 161 microvesicle-associated proteins, including many associated with the complement and coagulation signal-transduction cascades. Several proteases and protease inhibitors associated with acute phase responses were present, indicating that these microvesicles may be involved in these processes. There was a remarkably high variability in the protein content of plasma from different donors. In addition, we report that this variability could be relevant for their interaction with cellular systems. This work provides valuable information on plasma microvesicles and a foundation to understand microvesicle biology and clinical implications.