scholarly journals Effect of cortisol on the thiol content of rat liver nuclear proteins

1972 ◽  
Vol 126 (5) ◽  
pp. 1171-1179 ◽  
Author(s):  
D. Doenecke ◽  
M. Beato ◽  
L. F. Congote ◽  
C. E. Sekeris
Keyword(s):  
Rat Liver ◽  
Rna Synthesis ◽  
Soluble Fraction ◽  
Polyacrylamide Gel Electrophoresis ◽  
Nuclear Proteins ◽  
Carboxymethyl Lysine ◽  
Thiol Content ◽  
Liver Nuclei

Administration of cortisol to normal or adrenalectomized rats leads within 15–30min to an increased thiol content of nuclear proteins, measured by the incorporation of iodo[3H]-acetate or N-[14C]ethylmaleimide or by colorimetric methods. The same effect is observed after incubation of isolated rat liver nuclei with corticosteroids. The increased thiol content of the nuclear proteins shows the same time-dependence as the stimulation of RNA synthesis by corticosteroids observed in vivo and in vitro. Amino acid analysis of the carboxymethylated proteins reveals that in the experiments in vivo most of the label is present as carboxymethylcysteine with small amounts of carboxymethyl-lysine and carboxymethylhistidine, whereas in the experiments in vitro more carboxymethyl-lysine and carboxymethylhistidine than carboxymethylcysteine are found. The increase in the content of thiol groups is due to cleavage of the disulphide bridges between the nuclear proteins. Polyacrylamide-gel electrophoresis of the acid-soluble fraction reveals that most of the iodo[3H]acetate label is incorporated into a non-histone fraction with a molecular weight of approx. 45000 whereas in the acid-insoluble fractions many protein bands are labelled.

In vitro transfer of N,N′-diacetylchitobiose to glycoproteins in rat liver nuclei

1992 ◽  
Vol 70 (8) ◽  
pp. 677-683 ◽  
Author(s):  
Jacques Frot—Coutaz ◽  
Agnès Degiuli ◽  
Marie-Bénédicte Martel ◽  
Robert Létoublon
Keyword(s):  
Rat Liver ◽  
Biological Significance ◽  
Nuclear Proteins ◽  
Direct Transfer ◽  
Rat Liver Nuclei ◽  
Liver Nuclei

This work demonstrates that (N-acetyl[14C]glucosamine)2 is transferred from dolichyl pyrophosphate – (N-acetyl[14C]glucosamine)2 to endogenous nuclear glycoproteins. The (N-acetyl[l4C]gIucosamine)2 moiety is N-linked, since it can be released from the tryptic glycopeptides by N-glycosidase F and by hydrazinolysis, but not by β-elimination. The biological significance of this direct transfer of N, N′-diacetylchitobiose to nuclear proteins remains to be elucidated.Key words: nucleus, nuclear glycoproteins, nuclear N-acetylglucosaminyltransferase, N,N′ -diacetylchitobiose.

scholarly journals Role of membrane-bound and free polyribosomes in the synthesis of cytochrome c in rat liver

1974 ◽  
Vol 140 (2) ◽  
pp. 157-167 ◽  
Author(s):  
Néstor F. González-Cadavid ◽  
Carmen Sáez De Córdova
Keyword(s):  
Endoplasmic Reticulum ◽  
Cytochrome C ◽  
Rat Liver ◽  
Cell Structure ◽  
Polyacrylamide Gel Electrophoresis ◽  
Mitochondrial Protein ◽  
Sodium Dodecyl ◽  
Membrane Bound

The functional distinction of membrane-bound and free polyribosomes for the synthesis of exportable and non-exportable proteins respectively is not so strict as was initially thought, and it was therefore decided to investigate their relative contribution to the elaboration of an internal protein integrated into a cell structure. Cytochrome c was chosen as an example of a soluble mitochondrial protein, and the incorporation of [14C]leucine and δ-amino[14C]laevulinate into the molecule was studied by using different ribosomal preparations from regenerating rat liver. A new procedure was devised for the purification of cytochrome c, based on ion-exchange chromatography combined with sodium dodecyl sulphate–polyacrylamide-gel electrophoresis. In spite of cytochrome c being a non-exportable protein, the membrane-bound polyribosomes were at least as active as the free ribosomes in the synthesis in vitro of the apoprotein and the haem moiety. The detergent-treated ribosomes could also effect the synthesis of cytochrome c, although at a lower rate. Since in liver more than two-thirds of the ribosomes are bound to the endoplasmic-reticulum membranes, it is considered that in vivo they are responsible for the synthesis of most of the cytochrome c content of the cell. This suggests that in secretory tissues the endoplasmic reticulum plays a predominant role in mitochondrial biogenesis, although free ribosomes may participate in the partial turnover of some parts of the organelle. The hypothesis on the functional specialization of the different kinds of ribosomes was therefore modified to account for their parallel intervention in the synthesis of proteins associated with membranous structures.

scholarly journals Nuclear-envelope vesicles as a model system to study nucleocytoplasmic transport. Specific uptake of nuclear proteins

1987 ◽  
Vol 241 (1) ◽  
pp. 213-219 ◽  
Author(s):  
N Riedel ◽  
H Fasold
Keyword(s):  
Nuclear Envelope ◽  
Rat Liver ◽  
Protein Translocation ◽  
High Mobility ◽  
Preceding Paper ◽  
Nucleocytoplasmic Transport ◽  
Nuclear Proteins ◽  
Rat Liver Nuclei ◽  
Liver Nuclei

In the preceding paper [Riedel & Fasold (1987) Biochem. J. 241, 203-212] we have described a procedure for the preparation of nuclear-envelope vesicles (NE vesicles) from rat liver nuclei. These vesicles, which are largely free of components of the nuclear interior, were employed in an assay system in vitro to study protein translocation across the NE. We found that nuclear proteins such as histones, high-mobility-group proteins and acidic chromosomal proteins are specifically taken up and accumulated in the NE vesicles, whereas there is little or no affinity for non-nuclear proteins like immunoglobulin, myoglobin and cytochrome c. The kinetics of histone uptake into the NE vesicles are similar to those obtained for whole rat liver nuclei, and comparative studies with non-vesicular NEs prepared by deoxyribonuclease I-treatment (DNAase-NEs) indicate that the NE of the vesicles affects the uptake kinetics and increases the capacity for nuclear proteins. The uptake of histones into NE vesicles, but not the binding to DNAase-NEs, can be stimulated by GTP and GDP. Furthermore, we found that even very large molecules can be entrapped in the vesicles during their preparation. These results indicate that the NE vesicles might provide a useful system in vitro with which to investigate the structures and mechanisms involved in protein translocation across the NE.

Isolation and Analysis of Non-Histone Chromatin Proteins from Rat-Liver Nuclei by Three Different Methods

1974 ◽  
Vol 52 (12) ◽  
pp. 1143-1153 ◽  
Author(s):  
D. Suria ◽  
C. C. Liew
Keyword(s):  
Rat Liver ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Soluble Proteins ◽  
Polyacrylamide Gel ◽  
Pulse Labelling ◽  
Rat Liver Nuclei ◽  
Liver Nuclei ◽  
Chromatin Proteins

Non-histone chromatin proteins were isolated from rat-liver nuclei by three different methods, and defined as (I) phenol-soluble proteins, (II) SDS-soluble proteins and (III) proteins not adsorbed by cation-exchange chromatography. About 62–70% of chromatin proteins were recovered from the total nuclear proteins. The yield of non-histone chromatin proteins varied from 17 to 26% of chromatin proteins, depending on the method used. The amino-acid composition of these proteins showed that they are acidic in nature. Their phosphorus content was found to be 0.9, 1.1, and 1.4%, respectively, according to method I, II, or III. In-vivo pulse-labelling experiments indicated that chromatin proteins were highly labelled with 3H-acetate and 32P-phosphoric acid. In particular, the specific activities of 32P incorporation were higher in all non-histone chromatin proteins isolated as compared with histones. One-dimensional SDS–polyacrylamide gel electrophoresis showed that at least 26 similar fractions can be detected in the samples prepared by these three methods.The similarity of some of the proteins obtained from methods I and III was further confirmed by fractionation of the non-histone chromatin proteins in an isoelectro-focusing system followed by a second-dimensional SDS–polyacrylamide gel electrophoresis. It was found that more than 100 components could be identified. However, some minor variations of the non-histone chromatin proteins were detected by this system. The differences in proteins isolated by these methods are mainly quantitative rather than qualitative. The methods examined are not specific for the fractionation of a certain class of non-histone chromatin proteins.

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