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E K Davydova

Publications and source records attributed to E K Davydova.

At least 19 recordsLinked to original sources
PubMed2002

The phage N4 virion RNA polymerase catalytic domain is related to single-subunit RNA polymerases.

In vitro, bacteriophage N4 virion RNA polymerase (vRNAP) recognizes in vivo sites of transcription initiation on single-stranded templates. N4 vRNAP promoters are comprised of a hairpin structure and conserved sequences. Here, we show that vRNAP consists of a single 3500 amino acid polypeptide, and we define and characterize a transcriptionally active 1106 amino acid domain (mini-vRNAP). Biochemical and genetic characterization of this domain indicates that, despite its peculiar promoter specificity and lack of extensive sequence similarity to other DNA-dependent RNA polymerases, mini-vRNAP is related to the family of T7-like RNA polymerases.

Amino Acid Motifs↗
PubMed1998

The major core protein of messenger ribonucleoprotein particles (p50) promotes initiation of protein biosynthesis in vitro.

The major core protein of cytoplasmic messenger ribonucleoprotein particles (p50) has been shown previously to inhibit protein synthesis in vitro and in vivo. Furthermore, p50 is highly homologous to the Y-box-binding transcription factor family of proteins, binds DNA containing the Y-box motif, and thus may have a dual function in cells as a regulator of both transcription and translation. Here we show that binding or removal of p50 from rabbit reticulocyte lysate by monospecific antibodies to p50 strongly inhibits translation of endogenous and exogenous globin mRNAs as well as prokaryotic beta-galactosidase mRNA in a rabbit reticulocyte cell-free system. Thus, depending on the conditions, p50 not only may act as a translational repressor, but may also be required for protein synthesis. Translation inhibition with anti-p50 antibodies is not a result of mRNA degradation or its functional inactivation. The inhibition does not change the ribosome transit time, and therefore, it does not affect elongation/termination of polypeptide chains. The inhibition with anti-p50 antibodies is followed by a decay of polysomes and accumulation of the 48 S preinitiation complex. These results suggest that p50 participates in initiation of protein biosynthesis. Although uninvolved in the formation of the 48 S preinitiation complex, p50 is necessary either for attachment of the 60 S ribosomal subunit or for previous 5'-untranslated region scanning by the 43 S preinitiation complex.

Animals↗
PubMed1997

Overexpression in COS cells of p50, the major core protein associated with mRNA, results in translation inhibition.

p50, the major core protein of messenger ribonucleoprotein particles (mRNPs) in the cytoplasm of somatic mammalian cells, has been characterized previously as a member of the Y-box binding transcription factor family of proteins (YB-protein) by both high structural homology and ability to bind specifically the Y-box sequence in double-stranded DNA. YB proteins are present in a whole range of cell types and some have been identified as germ-specific cytoplasmic proteins masking stored mRNA from translation. Western blot analysis of the distribution of p50 in subcellular fractions of COS-1 cells shows that p50 is a cytoplasmic protein quantitatively associated with mRNA, both in polyribosomes and in free mRNPs. The level of p50 in COS-1 cells determined by Western immunoblotting is 0.10% of total protein, which is nearly equimolar to that of ribosomes and is approximately 5-10-fold higher than the mRNA level. Transient transfection of COS-1 cells with a p50-expressing vector results in a dramatic inhibition of protein synthesis. A control transfection with a vector expressing a frameshift mutant of p50 does not cause translation inhibition. Therefore the increase in p50 protein level is responsible for the inhibitory effect in these cells.

Animals↗
PubMed1993

Ribosomes terminated in vitro are in a tight association with non-phosphorylated elongation factor 2 (eEF-2) and GDP.

A proportion of the ribosome population in the eukaryotic cell is present in the form of single 80-S ribosomes. These are not involved in translation and are tightly associated with eukaryotic elongation factor 2 (eEF-2). The factor dissociates from ribosomes when it is ADP-ribosylated. Attempts at reconstitution of such complexes from ribosomal subunits and eEF-2 were not successful. We have shown that monomeric ribosomes in a tight complex with eEF-2 can be obtained in vitro as terminated ribosomes in a reconstituted translation system containing isolated polyribosomes, elongation factors and pH5 enzymes (all from rabbit reticulocytes). Incubation of the system with radioactive GTP demonstrated that terminated ribosomes contain GDP. ADP-ribosylation of eEF-2 bound to terminated ribosomes by diphtheria toxin leads to dissociation of both eEF-2 and GDP to the same extent. Thus the presence of GDP in terminated ribosomes is eEF-2 dependent. Ribosomes terminated in vitro as well as native single ribosomes contain the non-phosphorylated form of eEF-2. We assume that tight association of terminated ribosomes with the non-phosphorylated form of eEF-2 excludes both the ribosome and active eEF-2 from the translational cycle and thus, maintains the optimal proportion of translating ribosomes and free eEF-2 in the cell.

Animals↗
PubMed1990

ADP-ribosylated elongation factor 2 (ADP-ribosyl-EF-2) is unable to promote translocation within the ribosome.

Elongation factor 2 (EF-2), ADP-ribosylated in vitro by the A-fragment of diphtheria toxin, can (in the presence of GMPPCP) form stable complexes with ribosomes regardless of whether the ribosomes are empty or carrying poly(U) and Phe-tRNA in the A-site. Despite its efficient binding to ribosomes, ADP-ribosyl-EF-2, in contrast to the non-modified EF-2 is unable to promote the shift of Phe-tRNA from the A-site to the P-site of the ribosome as determined by the puromycin reaction, i.e. it is incapable of promoting the translocation reaction within the ribosome.

Adenosine Diphosphate Ribose↗
PubMed1989

Mechanism of elongation factor 2 (EF-2) inactivation upon phosphorylation. Phosphorylated EF-2 is unable to catalyze translocation.

Previously we have found that elongation factor 2 (EF-2) from mammalian cells can be phosphorylated by a special Ca2+/calmodulin-dependent protein kinase (EF-2 kinase). Phosphorylation results in complete inactivation of EF-2 in the poly(U)-directed cell-free translation system. However, the partial function of EF-2 affected by phosphorylation remained unknown. Here we show that phosphorylated EF-2, unlike non-phosphorylated EF-2, is unable to switch ribosomes carrying poly(U) and Phe-tRNA in the A site to a puromycin-reactive state. Thus, phosphorylation of EF-2 seems to block its ability to promote a shift of the aminoacyl(peptidyl)-tRNA from the A site to the P site, i.e. translocation itself.

Animals↗
PubMed1984

Phosphorylation of elongation factor 1 in polyribosome fraction of rabbit reticulocytes.

A single protein, Mr approximately 50000, is shown to be phosphorylated during incubation of a mono- and polyribosome fraction of rabbit reticulocytes with [gamma-32P]ATP at a low ionic strength. This protein has been identified as the elongation factor 1 alpha (EF-1 alpha). The phosphorylated EF-1 alpha, in contrast to the unmodified factor, is not detected in complexes with mono- and polyribosomes. It is suggested that the phosphorylation of EF-1 alpha can result in its decompartmentation from polyribosomes and thus affect the rate of protein synthesis.

Animals↗
PubMed1984

Eukaryotic elongation factor 2 loses its non-specific affinity for RNA and leaves polyribosomes as a result of ADP-ribosylation.

ADP-ribosylation of rabbit reticulocyte elongation factor 2 (EF-2) catalyzed by the A fragment of diphtheria toxin leads to a loss of its non-specific affinity for RNA. The removal of the ADP-ribose residue from EF-2 in the reverse reaction with nicotinamide restores its affinity for RNA. ADP-ribosylation of EF-2 is accompanied by its dissociation from the complexes with mono- and polyribosomes detected in the rabbit reticulocyte lysate at low ionic strength. The loss of the non-specific affinity of EF-2 for RNA as a result of ADP-ribosylation and, as a consequence, its decompartmentation from polyribosomes is assumed to be a reason for the diphtheria toxin-induced inactivation of the factor in eukaryotic cells.

Adenosine Diphosphate Ribose↗
PubMed1984

Endogenous ADP-ribosylation of elongation factor 2 in polyribosome fraction of rabbit reticulocytes.

Several polypeptides of about 120, 96, 85, 60 and 38 kDa are shown to be radiolabeled during incubation of the mono- and polyribosome fraction of rabbit reticulocytes with [32P]NAD. Among them is a polypeptide coinciding with elongation factor 2 (EF-2) in its electrophoretic mobility in SDS-polyacrylamide gel. The addition of pure EF-2 to the polyribosome fraction results in an increase of the radioactive label in this polypeptide band. From this it is concluded that both endogenous and added EF-2 is ADP-ribosylated by an enzyme associated with polyribosomes. A possibility of regulation of protein synthesis through endogenous ADP-ribosylation in vivo is considered.

Adenosine Diphosphate Ribose↗
PubMed1982

[The bulk of tRNA in the extracts of animal cells is not complexed with proteins].

The bulk of tRNA from cytoplasmic cell extracts of rat liver sedimented in the postribosomal zone appears to be in a free state and is not complexed with proteins. Evidence for this assumption may be derived from the following experimental findings: i) the tRNA of the extract is not adsorbed on nitrocellulose filters under conditions of ribonucleoprotein sorption; ii) after formaldehyde fixation and contrifugation to equilibrium in CsCl or Cs2SO4 density gradients the tRNA of the extract is distributed as free tRNA; iii) the tRNA of the extract does not differ considerably from the deproteinated tRNA in sedimentation velocity.

Animals↗
PubMedDate not supplied

Role of the lateral and medial hypothalamus in the reproduction of the motoric reaction which is a signal during the development of classical conditioned reflexes.

It has been demonstrated in experiments on three dogs that test electrostimulation of the lateral hypothalamus reproduces the motoric reaction which is the signal stimulus during the development of classical alimentary CR (83%) and does not reproduce it during the development of classical defensive CR (93%). The test electrostimulation of the medial divisions of the hypothalamus, like the electrostimulation of the LH, reproduces the "signal" motoric reaction, but in a smaller percent of cases (62%), during the development of classical alimentary CR, and does not reproduce it during the development of classical defensive CR. The reproduction of the signal motoric reaction during the electrostimulation of the LH is associated with the activation of a backward conditioned connection from the motivational structures of the hypothalamus to the representation of the signal stimulus in the motor cortex.

Aggression↗
PubMedDate not supplied

Role of the lateral and medial hypothalamus in the reproduction of alimentary and defensive instrumental reactions.

It was established in experiments on two dogs that a test electrical stimulation of the lateral hypothalamus reproduces an instrumental movement in the case of a developed alimentary instrumental CR to a tone and does not reproduce it in the case of a developed acid defensive instrumental CR. In the case of an electrical stimulation of the medial hypothalamus, the instrumental movement is essentially not reproduced, either in the alimentary or the defensive situations. The appearance of the instrumental movement with electro-stimulation of the LH in the alimentary situation is associated with activation of the backward conditioned connection from the motivational "alimentary" center of the LH to the representation of this movement in the motor cortex.

Acoustic Stimulation↗
PubMedDate not supplied

Movement as a signal during classical conditioning.

Analysis of the available data and that of the author disclosed the peculiarities of motor reaction when used as a conditioned stimulus. The author's data showed that if signal value is attributed to a motor reaction (passive movement or movement evoked by the direct stimulation of the motor cortex), the changes of excitability in the motor cortex representation of the dog's leg depend on the biological sign of the reinforcing stimulus during classic conditioning. They also remained the same during instrumental conditioning and were opposite in sign, showed increased excitability in the food situation, and decreased excitability in the defense situation. Using the movement as a conditional stimulus, we managed to uncover the commonality between classic and instrumental conditioning. This enabled us to answer questions, discussed by Pavlov and Guthrie, which, it seems to us, had not been convincingly answered during their time.

Animals↗