Dehydrogenase Binding Sites Abolish the “Dark” Fraction of NADH: Implication for Metabolic Sensing via FLIM

The present knowledge of the three-dimensional structure of ribosomes is far too limited to enable a complete understanding of the various roles which ribosomes play in protein biosynthesis. The spatial arrangement of proteins and ribonuclec acids in ribosomes can be analysed in many ways. Determination of binding sites for individual proteins on ribonuclec acid and locations of the mutual positions of proteins on the ribosome using labeling with fluorescent dyes, cross-linking reagents, neutron-diffraction or antibodies against ribosomal proteins seem to be most successful approaches. Structure and function of ribosomes can be correlated be depleting the complete ribosomes of some proteins to the functionally inactive core and by subsequent partial reconstitution in order to regain active ribosomal particles.

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Electron Probe Analysis of Muscle

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100054583 ◽

1982 ◽

Vol 40 ◽

pp. 398-401

Author(s):

A. V. Somlyo ◽

H. Shuman ◽

A. P. Somlyo

Keyword(s):

Skeletal Muscle ◽

Smooth Muscle ◽

Sarcoplasmic Reticulum ◽

Resting State ◽

Binding Sites ◽

Electron Probe ◽

Frog Skeletal Muscle ◽

Electron Probe Analysis ◽

Probe Analysis

Electron probe analysis of frozen dried cryosections of frog skeletal muscle, rabbit vascular smooth muscle and of isolated, hyperpermeab1 e rabbit cardiac myocytes has been used to determine the composition of the cytoplasm and organelles in the resting state as well as during contraction. The concentration of elements within the organelles reflects the permeabilities of the organelle membranes to the cytoplasmic ions as well as binding sites. The measurements of [Ca] in the sarcoplasmic reticulum (SR) and mitochondria at rest and during contraction, have direct bearing on their role as release and/or storage sites for Ca in situ.

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Sites of Insulin Action Using Ferritin Labeling

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100066759 ◽

1971 ◽

Vol 29 ◽

pp. 540-541

Author(s):

Burton B. Silver ◽

Ronald S. Nelson

Keyword(s):

Electron Microscopy ◽

Binding Sites ◽

Anterior Pituitary ◽

Insulin Action ◽

Diabetic Rats ◽

Insulin Antibody ◽

Fat Pad ◽

Target Organs ◽

Uranium Salts ◽

Sugar Levels

Some investigators feel that insulin does not enter cells but exerts its influence in some manner on the cell surface. Ferritin labeling of insulin and insulin antibody was used to determine if binding sites of insulin to specific target organs could be seen with electron microscopy.Alloxanized rats were considered diabetic if blood sugar levels were in excess of 300 mg %. Test reagents included ferritin, ferritin labeled insulin, and ferritin labeled insulin antibody. Target organs examined were were diaphragm, kidney, gastrocnemius, fat pad, liver and anterior pituitary. Reagents were administered through the left common carotid. Survival time was at least one hour in test animals. Tissue incubation studies were also done in normal as well as diabetic rats. Specimens were fixed in gluteraldehyde and osmium followed by staining with lead and uranium salts. Some tissues were not stained.

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Fluorescence ratio imaging of dynamic intracellular ionic signals

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100102298 ◽

1988 ◽

Vol 46 ◽

pp. 42-43

Author(s):

R. Y. Tsien ◽

A. Minta ◽

M. Poenie ◽

J.P.Y. Kao ◽

A. Harootunian

Keyword(s):

Binding Sites ◽

Living Cells ◽

Molecular Engineering ◽

Ph Indicators ◽

Highly Sensitive ◽

Fluorescence Ratio Imaging ◽

Ratio Imaging ◽

Technical Advances ◽

Indicator Dyes ◽

Fluorescein Dyes

Recent technical advances now enable the continuous imaging of important ionic signals inside individual living cells with micron spatial resolution and subsecond time resolution. This methodology relies on the molecular engineering of indicator dyes whose fluorescence is strong and highly sensitive to ions such as Ca2+, H+, or Na+, or Mg2+. The Ca2+ indicators, exemplified by fura-2 and indo-1, derive their high affinity (Kd near 200 nM) and selectivity for Ca2+ to a versatile tetracarboxylate binding site3 modeled on and isosteric with the well known chelator EGTA. The most commonly used pH indicators are fluorescein dyes (such as BCECF) modified to adjust their pKa's and improve their retention inside cells. Na+ indicators are crown ethers with cavity sizes chosen to select Na+ over K+: Mg2+ indicators use tricarboxylate binding sites truncated from those of the Ca2+ chelators, resulting in a more compact arrangement of carboxylates to suit the smaller ion.

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Lectin Binding Studies on RNA Tumor Virus-Infected Cells

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100115250 ◽

1975 ◽

Vol 33 ◽

pp. 326-327

Author(s):

D. C. Hixson

Keyword(s):

Binding Sites ◽

Lectin Binding ◽

Culture Conditions ◽

3T3 Cells ◽

Binding Studies ◽

Con A ◽

Microscopic Studies ◽

Tumor Virus ◽

Infected Cells ◽

Cell Culture Conditions

The abilities of plant lectins to preferentially agglutinate malignant cells and to bind to specific monosaccharide or oligosaccharide sequences of glycoproteins and glycolipids make them a new and important biochemical probe for investigating alterations in plasma membrane structure which may result from malignant transformation. Electron and light microscopic studies have demonstrated clustered binding sites on surfaces of SV40-infected or tryp- sinized 3T3 cells when labeled with concanavalin A (con A). No clustering of con A binding sites was observed in normal 3T3 cells. It has been proposed that topological rearrangement of lectin binding sites into clusters enables con A to agglutinate SV40-infected or trypsinized 3T3 cells (1). However, observations by other investigators have not been consistent with this proposal (2) perhaps due to differences in reagents used, cell culture conditions, or labeling techniques. The present work was undertaken to study the lectin binding properties of normal and RNA tumor virus-infected cells and their associated viruses using lectins and ferritin-conjugated lectins of five different specificities.

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