The Component Polypeptide Chains of Bovine Insulin Nucleate or Inhibit Aggregation of the Parent Protein in a Conformation-dependent Manner

2006 ◽  
Vol 360 (2) ◽  
pp. 497-509 ◽  
Author(s):  
Glyn L. Devlin ◽  
Tuomas P.J. Knowles ◽  
Adam Squires ◽  
Margaret G. McCammon ◽  
Sally L. Gras ◽  
...  
Keyword(s):  
Bovine Insulin ◽  
Dependent Manner ◽  
Parent Protein ◽  
Polypeptide Chains

Effects of insulin-like growth factor-I and insulin on the in-vitro uptake of sulphate by eel branchial cartilage: evidence for the presence of independent hepatic and pancreatic sulphation factors

1992 ◽  
Vol 133 (2) ◽  
pp. 211-219 ◽  
Author(s):  
C. Duan ◽  
T. Hirano
Keyword(s):  
Growth Factor ◽  
Coho Salmon ◽  
Bovine Insulin ◽  
Dependent Manner ◽  
Insulin Like Growth Factor ◽  
Cartilage Growth ◽  
Factor I ◽  
Igf I ◽  
Growth Factor I

ABSTRACT The possible roles of insulin-like growth factor-I (IGF-I) and insulin in regulating cartilage growth were studied in the teleost Anguilla japonica. Significant sulphation activity was found in the extracts of pancreas, liver and muscle, but not in those of kidney, intestine or spleen. The hepatic sulphation activity was significantly decreased by hypophysectomy or by fasting for 14 days, suggesting that this activity is regulated by pituitary function and nutritional status. Northern blot analysis revealed that the hepatic IGF-I mRNA in the eel consists of a major 4·0 kb band. This mRNA was GH-dependent and was significantly decreased by fasting for 14 days. On the other hand, fasting for 14 days had no significant effect on pancreatic sulphation activity. Pancreatic extracts from both intact and hypophysectomized eels exhibited equally significant stimulating activity. Addition of bovine or human insulin (1–250 ng/ml) to the culture medium significantly stimulated sulphate uptake in a dose-dependent manner. Teleost (coho salmon) insulin was as effective as bovine insulin. Bovine insulin was more effective than IGF-I at lower concentrations (1–4 ng/ml) but less effective at higher concentrations (10–250 ng/ml). These results indicate that not only IGF-I but also insulin are likely to be involved in the regulation of cartilage growth in the eel. Journal of Endocrinology (1992) 133, 211–219

Effects of growth hormone and triiodothyronine on insulin-induced progesterone production by granulosa cells from prepubertal gilts

1992 ◽  
Vol 72 (3) ◽  
pp. 589-593 ◽  
Author(s):  
R. N. Kirkwood ◽  
P. A. Thacker ◽  
K. Rajkumar
Keyword(s):  
Growth Hormone ◽  
Granulosa Cells ◽  
Culture Medium ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Bovine Insulin ◽  
Dependent Manner ◽  
Progesterone Production ◽  
Porcine Granulosa Cells

Two experiments were performed using granulosa cells from medium-sized follicles (2–4 mm) derived from prepubertal gilts. Cells were cultured in a serum-free medium at a density of either 1 or 2 × 106 viable cells per well (experiments 1 and 2, respectively). For exp. 1, porcine growth hormone (pGH) (0 or 100 ng mL−1) was included in the culture medium from the time of plating, and low-density lipoprotein (LDL) (100 μg mL−1) was added at 72 h. For exp. 2, granulosa cells were plated in a culture medium containing either pGH (0 or 100 ng mL−1) or triiodothyronine (T3) (0 or 5 ng mL−1) or both pGH T3; LDL was not included. For both experiments, after 24 h of culture, bovine insulin at 0, 10, 100 or 1000 ng mL−1 was included in the medium. Hormones were replaced at 48 and 72 h, and the cultures were terminated at 96 h. Results from exp. 1 indicated that insulin increased (P < 0.01) progesterone production in a dose-dependent manner, both in the presence and absence of LDL. This response was augmented (P < 0.01) by co-culture with pGH. Results from exp. 2 confirmed the augmenting effect of pGH (P < 0.01). It was further observed that T3 increased (P < 0.01) progesterone production when cultured with insulin at 1000 ng mL−1, but at lower insulin-inclusion levels, results were equivocal. The progesterone production response was greatest (P < 0.01) when cells were cultured with both pGH and T3 at insulin levels of 100 or 1000 ng mL−1. There appeared to be little relationship between the media concentrations of insulin-like growth factor 1 and progesterone. The present results suggest that relatively high levels of pGH and T3 will enhance the in vitro steroidogenic capabilities of porcine granulosa cells. Key words: Granulosa cells, GH, T3, insulin

Eel insulin: isolation, characterization and stimulatory actions on [35S]sulphate and [3H]thymidine uptake in the branchial cartilage of the eel in vitro

1992 ◽  
Vol 133 (2) ◽  
pp. 221-230 ◽  
Author(s):  
C. Duan ◽  
T. Noso ◽  
S. Moriyama ◽  
H. Kawauchi ◽  
T. Hirano
Keyword(s):  
Bovine Insulin ◽  
Cartilage Matrix ◽  
Common Mechanism ◽  
Thymidine Uptake ◽  
Dependent Manner ◽  
Matrix Synthesis ◽  
Sulphate Uptake ◽  
Igf I ◽  
Dose Dependent

ABSTRACT Our previous studies have shown that mammalian and salmon insulins stimulate sulphate uptake by cultured eel cartilage, suggesting the possible involvement of insulin in the regulation of cartilage matrix synthesis. In the present study, homologous eel insulin was isolated and characterized, and its effects on cartilage matrix synthesis and DNA synthesis were examined in vitro. Insulin was extracted from eel pancreas with acid–ethanol, and subsequently purified by isoelectric precipitation at pH 5·3, gel filtration on Sephadex G-50, and reversed-phase high-performance liquid chromatography. The amino acid composition and complete sequence (50 residues) of eel insulin revealed high homology to teleostean and mammalian insulins. The isolated eel insulin produced a more pronounced and longer lasting hypoglycaemic effect than bovine insulin in the eel. Homologous eel insulin, like bovine insulin-like growth factor (IGF-I) and insulin, stimulated sulphate uptake by cultured eel cartilage in a dose-dependent manner (16–1000 ng/ml). Combination experiments using maximal concentrations of bovine IGF-I (250 ng/ml) and increasing amounts of eel insulin (10–250 ng/ml) showed no additive effects of insulin on sulphate uptake, suggesting that insulin and IGF-I may share a common mechanism(s) of action. Eel insulin and bovine IGF-I also enhanced thymidine incorporation by eel cartilage in a dose-dependent manner (4–1000 ng/ml); eel insulin was equipotent with bovine IGF-I. These results suggest that insulin, like IGF-I, may exert direct growth-promoting actions in branchial cartilage of the eel. Journal of Endocrinology (1992) 133, 221–230

The conformation of an atypical IgG myeloma protein and its papain fragments

1970 ◽  
Vol 48 (7) ◽  
pp. 784-789 ◽  
Author(s):  
G. E. Connell ◽  
K. J. Dorrington ◽  
A. F. Lewis ◽  
D. M. Parr
Keyword(s):  
Molecular Weight ◽  
The Other ◽  
Optical Rotatory Dispersion ◽  
Rotatory Dispersion ◽  
Optical Rotatory ◽  
Fab Fragment ◽  
Myeloma Protein ◽  
Myeloma Proteins ◽  
Parent Protein ◽  
Polypeptide Chains

An immunoglobulin IgG (Sackfield) which is known to have polypeptide chains shorter than those of typical proteins of its class has been subjected to fragmentation by papain in the presence of cysteine. One fragment was recovered which was indistinguishable from normal Fc fragment. The other fragment was related to normal Fab fragment but differed from it in several of its properties. The molecular weight was only one-half that of normal Fab. The optical rotatory dispersion spectrum of IgG (Sackfield) had features which differed from those of typical IgG myeloma proteins. The optical rotatory dispersion spectrum of Fc (Sackfield) was identical with those of other myeloma proteins, while the Fab (Sackfield) spectrum reflected the differences observed in the parent protein.

Antibodies against the Component Polypeptide Chains of Bovine Insulin

Science ◽  
1965 ◽  
Vol 147 (3658) ◽  
pp. 617-619 ◽  
Author(s):  
Y. Yagi ◽  
P. Maier ◽  
D. Pressman
Keyword(s):  
Bovine Insulin ◽  
Polypeptide Chains

scholarly journals Hypochlorite-induced oxidation of proteins in plasma: formation of chloramines and nitrogen-centred radicals and their role in protein fragmentation

1999 ◽  
Vol 340 (2) ◽  
pp. 539-548 ◽  
Author(s):  
Clare L. HAWKINS ◽  
Michael J. DAVIES
Keyword(s):  
Plasma Protein ◽  
Plasma Proteins ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Nitrobenzoic Acid ◽  
Parent Protein ◽  
Sds Page ◽  
Amine Groups ◽  
Epr Spin Trapping ◽  
Protein Fragmentation

Activated phagocyte cells generate hypochlorite (HOCl) via the release of H2O2 and the enzyme myeloperoxidase. Plasma proteins are major targets for HOCl, although little information is available about the mechanism(s) of oxidation. In this study the reaction of HOCl (at least 50 μM) with diluted fresh human plasma has been shown to generate material that oxidizes 5-thio-2-nitrobenzoic acid; these oxidants are believed to be chloramines formed from the reaction of HOCl with protein amine groups. Chloramines have also been detected with isolated plasma proteins treated with HOCl. In both cases chloramine formation accounts for approx. 20-30% of the added HOCl. These chloramines decompose in a time-dependent manner when incubated at 20 or 37 °C but not at 4 °C. Ascorbate and urate remove these chloramines in a time- and concentration-dependent manner, with the former being more efficient. The reaction of fresh diluted plasma with HOCl also gives rise to protein-derived nitrogen-centred radicals in a time- and HOCl-concentration-dependent manner; these have been detected by EPR spin trapping. Identical radicals have been detected with isolated HOCl-treated plasma proteins. Radical formation was inhibited by excess methionine, implicating protein-derived chloramines (probably from lysine side chains) as the radical source. Plasma protein fragmentation occurs in a time- and HOCl-concentration-dependent manner, as evidenced by the increased mobility of the EPR spin adducts, the detection of further radical species believed to be intermediates in protein degradation and the loss of the parent protein bands on SDS/PAGE. Fragmentation can be inhibited by methionine and other agents (ascorbate, urate, Trolox C or GSH) capable of removing chloramines and reactive radicals. These results are consistent with protein-derived chloramines, and the radicals derived from them, as contributing agents in HOCl-induced plasma protein oxidation.

Immunogold labeling of the calcium binding protein synexin in isolated adrenal chromaffin granules and chromaffin cells

1990 ◽  
Vol 48 (3) ◽  
pp. 952-953
Author(s):  
Gemma A.J. Kuijpers ◽  
Harvey B. Pollard
Keyword(s):  
Adrenal Medulla ◽  
Calcium Binding ◽  
Chromaffin Cells ◽  
Cell Activation ◽  
Structural Information ◽  
Dependent Manner ◽  
Chromaffin Granules ◽  
Immuno Electron Microscopy ◽  
Ultrathin Sections ◽  
Adrenal Chromaffin

Exocytotic fusion of granules in the adrenal medulla chromaffin cell is triggered by a rise in the concentration of cytosolic Ca2+ upon cell activation. The protein synexin, annexin VII, was originally found in the adrenal medulla and has been shown to cause aggregation and to support fusion of chromaffin granules in a Ca2+-dependent manner. We have previously suggested that synexin may there fore play a role in the exocytotic fusion process. In order to obtain more structural information on synexin, we performed immuno-electron microscopy on frozen ultrathin sections of both isolated chromaffin granules and chromaffin cells.Chromaffin granules were isolated from bovine adrenal medulla, and synexin was isolated from bovine lung. Granules were incubated in the presence or absence of synexin (24 μg per mg granule protein) and Ca2+ (1 mM), which induces maximal granule aggregation, in 0.3M sucrose-40m MMES buffer(pH 6.0). Granules were pelleted, washed twice in buffer without synexin and fixed with 2% glutaraldehyde- 2% para formaldehyde in 0.1 M phosphate buffer (GA/PFA) for 30 min. Chromaffin cells were isolated and cultured for 3-5 days, and washed and incubated in Krebs solution with or without 20 uM nicotine. Cells were fixed 90 sec after on set of stimulation with GA/PFA for 30 min. Fixed granule or cell pellets were washed, infiltrated with 2.3 M sucrose in PBS, mounted and frozen in liquid N2.

Insights into amyloid disease from fly models

2014 ◽  
Vol 56 ◽  
pp. 69-83 ◽  
Author(s):  
Ko-Fan Chen ◽  
Damian C. Crowther
Keyword(s):  
Drosophila Melanogaster ◽  
Neurodegenerative Disorders ◽  
Amyloid Β ◽  
Amyloid Β Peptide ◽  
Disease Pathogenesis ◽  
Amyloid Aggregates ◽  
Aβ Amyloid ◽  
Polypeptide Chains ◽  
Amyloid Diseases

The formation of amyloid aggregates is a feature of most, if not all, polypeptide chains. In vivo modelling of this process has been undertaken in the fruitfly Drosophila melanogaster with remarkable success. Models of both neurological and systemic amyloid diseases have been generated and have informed our understanding of disease pathogenesis in two main ways. First, the toxic amyloid species have been at least partially characterized, for example in the case of the Aβ (amyloid β-peptide) associated with Alzheimer's disease. Secondly, the genetic underpinning of model disease-linked phenotypes has been characterized for a number of neurodegenerative disorders. The current challenge is to integrate our understanding of disease-linked processes in the fly with our growing knowledge of human disease, for the benefit of patients.

Spindle scaling mechanisms

2020 ◽  
Vol 64 (2) ◽  
pp. 383-396
Author(s):  
Lara K. Krüger ◽  
Phong T. Tran
Keyword(s):  
Cell Size ◽  
Spindle Assembly ◽  
Dependent Manner ◽  
Post Translational Modification ◽  
Size Dependent ◽  
A Cell ◽  
Chromosome Separation ◽  
Spindle Elongation ◽  
Protein Gradients ◽  
Spindle Structure

Abstract The mitotic spindle robustly scales with cell size in a plethora of different organisms. During development and throughout evolution, the spindle adjusts to cell size in metazoans and yeast in order to ensure faithful chromosome separation. Spindle adjustment to cell size occurs by the scaling of spindle length, spindle shape and the velocity of spindle assembly and elongation. Different mechanisms, depending on spindle structure and organism, account for these scaling relationships. The limited availability of critical spindle components, protein gradients, sequestration of spindle components, or post-translational modification and differential expression levels have been implicated in the regulation of spindle length and the spindle assembly/elongation velocity in a cell size-dependent manner. In this review, we will discuss the phenomenon and mechanisms of spindle length, spindle shape and spindle elongation velocity scaling with cell size.

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