Genetic dissection of the final exocytosis steps in Paramecium tetraurelia cells: cytochemical determination of Ca2+-ATPase activity over performed exocytosis sites

1980 ◽  
Vol 46 (1) ◽  
pp. 17-40 ◽  
Author(s):  
H. Plattner ◽  
K. Reichel ◽  
H. Matt ◽  
J. Beisson ◽  
M. Lefort-Tran ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Freeze Fracture ◽  
Secretory Vesicles ◽  
Genetic Dissection ◽  
Paramecium Tetraurelia ◽  
Electron Microscopic ◽  
X Ray ◽  
Electron Microscopic Cytochemistry ◽  
Function Correlation

In different Paramecium tetraurelia strains the occurrence of a Ca2+-ATPase (or p-nitro-phenylphosphatase) activity at the preformed attachment and exocytosis sites of the secretory vesicles (trichocysts) was analysed by electron-microscopic cytochemistry and X-ray microanalysis. In conjunction with freeze-fracture studies it was found that only those strains, which contain rosette particles, display this Ca2+-ATPase activity (7S, K 40I, nd 9 (18 degrees C), while other strains (nd 6, nd 9 (27 degrees C), tam 38) are devoid of both these characteristics. The presence (absence) of rosette particles and of Ca2+-ATPase activity at the preformed exocytosis sites is correlated with the capability (incapability) of performing exocytosis in these strains. We discuss several possible interpretations of this structure-function correlation.

Direct visualization of phase-separated domains in lipid membranes

1978 ◽  
Vol 36 (2) ◽  
pp. 290-291
Author(s):  
S. W. Hui ◽  
T. P. Stewart
Keyword(s):  
Lipid Membranes ◽  
Structural Information ◽  
Freeze Fracture ◽  
Biological Molecules ◽  
Vacuum Drying ◽  
Direct Visualization ◽  
X Ray Diffraction ◽  
Electron Microscopic ◽  
X Ray ◽  
Hydrocarbon Chains

Direct electron microscopic study of biological molecules has been hampered by such factors as radiation damage, lack of contrast and vacuum drying. In certain cases, however, the difficulties may be overcome by using redundent structural information from repeating units and by various specimen preservation methods. With bilayers of phospholipids in which both the solid and fluid phases co-exist, the ordering of the hydrocarbon chains may be utilized to form diffraction contrast images. Domains of different molecular packings may be recgnizable by placing properly chosen filters in the diffraction plane. These domains would correspond to those observed by freeze fracture, if certain distinctive undulating patterns are associated with certain molecular packing, as suggested by X-ray diffraction studies. By using an environmental stage, we were able to directly observe these domains in bilayers of mixed phospholipids at various temperatures at which their phases change from misible to inmissible states.

Polarity Determination in Sphalerite and Wurtzite Structures

1990 ◽  
Vol 48 (2) ◽  
pp. 500-501
Author(s):  
Stuart McKernan ◽  
C. Barry Carter
Keyword(s):  
Opposite Polarity ◽  
Single Domain ◽  
Polar Material ◽  
Electron Microscopic ◽  
Dynamical Interaction ◽  
X Ray ◽  
Polarity Determination ◽  
The Absolute ◽  
Microscopic Techniques

The determination of the absolute polarity of a polar material is often crucial to the understanding of the defects which occur in such materials. Several methods exist by which this determination may be performed. In bulk, single-domain specimens, macroscopic techniques may be used, such as the different etching behavior, using the appropriate etchant, of surfaces with opposite polarity. X-ray measurements under conditions where Friedel’s law (which means that the intensity of reflections from planes of opposite polarity are indistinguishable) breaks down can also be used to determine the absolute polarity of bulk, single-domain specimens. On the microscopic scale, and particularly where antiphase boundaries (APBs), which separate regions of opposite polarity exist, electron microscopic techniques must be employed. Two techniques are commonly practised; the first [1], involves the dynamical interaction of hoLz lines which interfere constructively or destructively with the zero order reflection, depending on the crystal polarity. The crystal polarity can therefore be directly deduced from the relative intensity of these interactions.

Electron Microscopic Evidence for the Transmembrane Displa cement of Calcium ATPase

1984 ◽  
Vol 39 (1-2) ◽  
pp. 177-179 ◽  
Author(s):  
Donald J. Scales ◽  
Stefan R. Highsmith
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Atpase Activity ◽  
Freeze Fracture ◽  
Calcium Atpase ◽  
Two Dimensional ◽  
Electron Microscopic ◽  
Microscopic Evidence ◽  
Sarcoplasmic Reticulum Membrane ◽  
First Time

Abstract Incubation of the Ca2+-ATPase in vanadate solutions leads to the formation of two-dimensional arrays in the sarcoplasmic reticulum membrane. Electron micrographic freeze fracture replicas show depressions on the inner leaflet for the first time. This indicates that the ATPase has moved perpendicular to the plane of the membrane. Our results also suggest that aggregation of the Ca2+-ATPase into the two-dimensional arrays occurs before they move into the membrane. These phenomena were observed as soon as 15 minutes after vanadate was added. The effects of vanadate appear to be completely reversible. When SR was incubated in the vanadate solutions and was then diluted into a buffer containing Ca2+ and ATP, the ATPase activity was normal for up to several hours of incubation and only somewhat reduced after 3 days.

scholarly journals Preliminary X-ray crystallographic analysis of an engineered glutamyl-tRNA synthetase fromEscherichia coli

2014 ◽  
Vol 70 (7) ◽  
pp. 922-927 ◽  
Author(s):  
Nipa Chongdar ◽  
Saumya Dasgupta ◽  
Ajit Bikram Datta ◽  
Gautam Basu
Keyword(s):  
Structure Function ◽  
Thermus Thermophilus ◽  
Trna Synthetase ◽  
Crystallographic Analysis ◽  
X Ray ◽  
E Coli ◽  
Crystal Contacts ◽  
Function Correlation ◽  
Trna Substrate

The nature of interaction between glutamyl-tRNA synthetase (GluRS) and its tRNA substrate is unique in bacteria in that many bacterial GluRS are capable of recognizing two tRNA substrates: tRNAGluand tRNAGln. To properly understand this distinctive GluRS–tRNA interaction it is important to pursue detailed structure–function studies; however, because of the fact that tRNA–GluRS interaction in bacteria is also associated with phylum-specific idiosyncrasies, the structure–function correlation studies must also be phylum-specific. GluRS fromThermus thermophilusandEscherichia coli, which belong to evolutionarily distant phyla, are the biochemically best characterized. Of these, only the structure ofT. thermophilusGluRS is available. To fully unravel the subtleties of tRNAGlu–GluRS interaction inE. coli, a model bacterium that can also be pathogenic, determination of theE. coliGluRS structure is essential. However, previous attempts have failed to crystallizeE. coliGluRS. By mapping crystal contacts of a homologous GluRS onto theE. coliGluRS sequence, two surface residues were identified that might have been hindering crystallization attempts. Accordingly, these two residues were mutated and crystallization of the double mutant was attempted. Here, the design, expression, purification and crystallization of an engineeredE. coliGluRS in which two surface residues were mutated to optimize crystal contacts are reported.

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