In vitro labeling of proteins by reductive methylation: Application to proteins involved in supramolecular structures

1982 ◽  
Vol 19 (1) ◽  
pp. 77-91 ◽  
Author(s):  
C. S. Heacock ◽  
B. W. Bernstein ◽  
A. S. Duhaiman ◽  
D. A. Amorese ◽  
J. R. Bamburg
Keyword(s):  
Supramolecular Structures ◽  
Reductive Methylation

Absence of albumin receptor on brain capillaries in vivo or in vitro

1985 ◽  
Vol 249 (3) ◽  
pp. E264-E267 ◽  
Author(s):  
W. M. Pardridge ◽  
J. Eisenberg ◽  
W. T. Cefalu
Keyword(s):  
Transit Time ◽  
Bovine Brain ◽  
In Vitro Assays ◽  
Injection Technique ◽  
Receptor Model ◽  
Brain Capillaries ◽  
Reductive Methylation ◽  
Albumin Receptor

Hormones and drugs are known to be available for transport into brain and liver in vivo from the circulating albumin-bound pool. An albumin receptor-mediated mechanism is one possible way in which the transport of ligands from the circulating albumin-bound pool into the tissue may be catalyzed. The albumin receptor model was tested for brain in the present studies using both 125I-albumin (labeled by lactoperoxidase) and [3H]albumin (labeled by reductive methylation). The interaction of the labeled albumin with brain capillaries was assessed in vivo with the carotid injection technique in rats and in vitro with isolated bovine brain capillaries. Artifactually high nonspecific binding in both the in vivo and in vitro assays was observed with 125I-albumin. Conversely, the transit time of [3H]albumin through the brain capillaries in vivo was no greater than the transit time of [14C]sucrose. The binding of [3H]albumin to isolated microvessels in vitro was low, less than [3H]inulin and was nonsaturable. In conclusion these studies do not support the albumin receptor model for the transport of albumin-bound ligands into tissues such as brain.

scholarly journals Sound attenuation of polymerizing actin reflects supramolecular structures: viscoelastic properties of actin gels modified by cytochalasin D, profilin and α-actinin

2001 ◽  
Vol 355 (3) ◽  
pp. 771-778 ◽  
Author(s):  
Oliver WAGNER ◽  
Herwig SCHÜLER ◽  
Peter HOFMANN ◽  
David LANGER ◽  
Peter DANCKER ◽  
...  
Keyword(s):  
Sound Velocity ◽  
Actin Filaments ◽  
Actin Polymerization ◽  
Cytochalasin D ◽  
Supramolecular Structures ◽  
Lag Phase ◽  
Acoustic Attenuation ◽  
Ultrasound Attenuation ◽  
Actin Concentration

Polymerization and depolymerization of cytoskeletal elements maintaining cytoplasmic stiffness are key factors in the control of cell crawling. Rheometry is a significant tool in determining the mechanical properties of the single elements in vitro. Viscoelasticity of gels formed by these polymers strongly depends on both the length and the associations of the filaments (e.g. entanglements, annealings and side-by-side associations). Ultrasound attenuation is related to viscosity, sound velocity and supramolecular structures in the sample. In combination with a small glass fibre (2mm×50µm), serving as a viscosity sensor, an acoustic microscope was used to measure the elasticity and acoustic attenuation of actin solutions. Changes in acoustic attenuation of polymerizing actin by far exceed the values expected from calculations based on changes in viscosity and sound velocity. During the lag-phase of actin polymerization, attenuation slightly decreases, depending on actin concentration. After the half-maximum viscosity is accomplished and elasticity turns into steady state, attenuation distinctly rises. Changes in ultrasound attenuation depend on actin concentration, and they are modulated by the addition of α-actinin, cytochalasin D and profilin. Thus absorption and scattering of sound on the polymerization of actin is related to the packing density of the actin net, entanglements and the length of the actin filaments. Shortening of actin filaments by cytochalasin D was also confirmed by electron micrographs and falling-ball viscosimetry. In addition to viscosity and elasticity, the attenuation of sound proved to be a valuable parameter in characterizing actin polymerization and the supramolecular associations of F-actin.

CBG does not restrict blood-brain barrier corticosterone transport in rabbits

1986 ◽  
Vol 251 (2) ◽  
pp. E204-E208
Author(s):  
W. M. Pardridge ◽  
J. Eisenberg ◽  
G. Fierer ◽  
R. W. Kuhn
Keyword(s):  
Specific Binding ◽  
Rabbit Brain ◽  
Metabolic Clearance ◽  
Brain Capillaries ◽  
Reductive Methylation ◽  
Physiological Concentration ◽  
Peripheral Tissues ◽  
Corticosteroid Binding Globulin

The metabolic clearance rate of corticosterone in rabbits is unrelated to the physiological concentration of corticosteroid binding globulin (CBG) in rabbit plasma. This suggests that corticosterone is available for transport into peripheral tissues in rabbits from the circulating CBG-bound pool, similar to what is known to occur in rat liver. This hypothesis was tested in the present studies, which investigate the transport of corticosterone into rabbit brain from the circulating rabbit or human CBG-bound pool. Corticosterone was readily exchangeable in brain capillaries in vivo from the circulating albumin-bound and rabbit or human CBG-bound pools. The involvement of specific CBG receptors on brain capillary endothelia in this process was investigated with [3H]-labeled human CBG prepared by reductive methylation. The transport of [3H]CBG across rabbit brain capillaries in vivo was immeasurably low, and no specific binding of this radiolabeled plasma protein to isolated brain capillaries in vitro was observed at 37 degrees C during incubations up to 120 min. These studies indicate that the rabbit is a novel system for assessing the role of CBG in delivering corticosterone to peripheral tissues in vivo and that specific endothelial CBG receptors may not participate in the transport process.

scholarly journals Effects of ammonia concentration and microbial population on in vitro degradation of 14C-labelled dietary proteins

1991 ◽  
Vol 71 (1) ◽  
pp. 125-133
Author(s):  
M. K. Song ◽  
J. J. Kennelly
Keyword(s):  
Protein Degradation ◽  
Microbial Population ◽  
Rumen Fluid ◽  
Protein Solubility ◽  
Ammonia Concentration ◽  
Barley Grain ◽  
Minor Role ◽  
Reductive Methylation ◽  
Ruminal Fluid

Rumen fluid from two nonlactating cows fed barley silage and rolled barley grain based concentrates (75:25 on a dry matter basis) was incubated for 0.5, 1, 2, 3, and 4 h with 14C-labelled soybean meal (SBM), fish meal (FM) and corn gluten meal (CGM) to examine the effects of ammonia concentration and protein solubility on rate and extent of protein degradation by total mixed ruminal microorganisms (TMM) or mixed ruminal bacteria (MB). Proteins were labelled by reductive methylation. Ammonia concentration in control ruminal fluid was 4.0 mg dL−1; graded levels of 1 M (NH4)2SO4 were added to achieve ammonia concentrations of 10, 20, and 30 mg dL−1 ruminal fluid. Soluble protein was extracted with borate-phosphate buffer (pH 7.8). Removal of protozoa reduced the extent of protein degradation. However, relative to bacteria, protozoa appeared to play a minor role in protein degradation. Rate and extent of protein degradation was not influenced by ammonia concentration, but was highly correlated with protein solubility. Therefore, it is concluded that ammonia concentration is not the primary factor regulating proteolytic activity of rumen microorganisms, rather their activities may depend upon the solubility and physico-chemical properties of proteins. Key words: In vitro, ammonia concentration, protein degradation, microbial population

Directed Assembly of Elastin-like Proteins into defined Supramolecular Structures and Cargo Encapsulation In Vitro

10.3791/60935 ◽  
2020 ◽  
Author(s):  
Andreas Schreiber ◽  
Lara G. Stühn ◽  
Süreyya E. Geissinger ◽  
Matthias C. Huber ◽  
Stefan M. Schiller
Keyword(s):  
Directed Assembly ◽  
Supramolecular Structures

Ultrastructural Investigations of the Contractile Filaments of Amoebae

1974 ◽  
Vol 32 ◽  
pp. 188-189
Author(s):  
P.L. Moore
Keyword(s):  
Cytoplasmic Streaming ◽  
Three Dimensional ◽  
Freeze Fracture ◽  
Amoeboid Movement ◽  
Different Types ◽  
Chemical Conditions ◽  
Complete Interpretation

Previous freeze fracture results on the intact giant, amoeba Chaos carolinensis indicated the presence of a fibrillar arrangement of filaments within the cytoplasm. A complete interpretation of the three dimensional ultrastructure of these structures, and their possible role in amoeboid movement was not possible, since comparable results could not be obtained with conventional fixation of intact amoebae. Progress in interpreting the freeze fracture images of amoebae required a more thorough understanding of the different types of filaments present in amoebae, and of the ways in which they could be organized while remaining functional.The recent development of a calcium sensitive, demembranated, amoeboid model of Chaos carolinensis has made it possible to achieve a better understanding of such functional arrangements of amoeboid filaments. In these models the motility of demembranated cytoplasm can be controlled in vitro, and the chemical conditions necessary for contractility, and cytoplasmic streaming can be investigated. It is clear from these studies that “fibrils” exist in amoeboid models, and that they are capable of contracting along their length under conditions similar to those which cause contraction in vertebrate muscles.

In Vitro Induction of Crystals of MT Protein by Vinblastine-Colcemid in Mouse Spermatids

1974 ◽  
Vol 32 ◽  
pp. 132-133
Author(s):  
John J. Wolosewick ◽  
John H. D. Bryan
Keyword(s):  
Endoplasmic Reticulum ◽  
Smooth Endoplasmic Reticulum ◽  
Nuclear Ring ◽  
Rough Er ◽  
Distal Direction ◽  
In Vitro Induction

Early in spermiogenesis the manchette is rapidly assembled in a distal direction from the nuclear-ring-densities. The association of vesicles of smooth endoplasmic reticulum (SER) and the manchette microtubules (MTS) has been reported. In the mouse, osmophilic densities at the distal ends of the manchette are the organizing centers (MTOCS), and are associated with the SER. Rapid MT assembly and the lack of rough ER suggests that there is an existing pool of MT protein. Colcemid potentiates the reaction of vinblastine with tubulin and was used in this investigation to detect this protein.

Induction and Differentiation of Dental Epithelium in Vivo and in Vitro

1982 ◽  
Vol 40 ◽  
pp. 142-145
Author(s):  
E. J. Kollar
Keyword(s):  
Connective Tissue ◽  
Oral Mucosa ◽  
Basal Lamina ◽  
Functional Derivatives ◽  
Specific Source ◽  
Dental Epithelium ◽  
Connective Tissue Stroma

The differentiation and maintenance of many specialized epithelial structures are dependent on the underlying connective tissue stroma and on an intact basal lamina. These requirements are especially stringent in the development and maintenance of the skin and oral mucosa. The keratinization patterns of thin or thick cornified layers as well as the appearance of specialized functional derivatives such as hair and teeth can be correlated with the specific source of stroma which supports these differentiated expressions.

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