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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A system of translation of matrix-bound poly(U) by purified Escherichia coli ribosomes was used to obtain pre-translocation state ribosomes in columns and then to induce translocation under controlled conditions by passing the elongation factor G (EF-G) with the non-cleavable GTP analog (guanylyl-methylene diphosphonate). It has been shown that translocation in the ribosome, checked by the release of deacylated tRNA, as well as by the puromycin reaction, is induced by the attachment of EF-G (with the non-cleavable GTP analog) to the ribosome and not by its detachment. In accordance with this, the ionic conditions under which the affinity of EF-G with the GTP analog to the ribosome is increased (NH4Cl instead of KCl, a lowered ionic strength) have been also found to be more effective for translocation. On the other hand, it has been shown that the detachment (removal) of EF-G is a strict pre-requisite for the appearance of competence to bind the next aminoacyl-tRNA, and thus for a continuation of the elongation cycle. A conclusion is made that the mechanical shifts of products and substrates, such as peptidyl-tRNA and deacylated tRNA, within the ribosome in the process of translocation are promoted only by the affinity of EF-G to the ribosome and does not depend on the cleavage of GTP. On the basis of the results obtained, the following sequence of events is deduced for the process of EF-G-promoted translocation: 1) interaction of EF-G.GTP with the pre-translocative ribosome, 2) translocation displacements of products and substrates, including the release of deacylated tRNA (probably conjugated with the shift of mRNA), 3) GTP hydrolysis, 4) release of EF-G and GTP from the post-translocated ribosome.
Mol Biol (Mosk)
PMID:[Sequence of events in the process of factor-promoted translocation in the ribosome]. 35 74

The kinetic and regulatory properties of purified rat heart AMP deaminase were investigated. In the presence of 100 mM KCl, the enzyme exhibited a slightly sigmoid-shaped plot of reaction rate, vs. substrate concentration, which shifted to a more hyperbolic form when ATP, ADP or GTP were added. ATP was the most potent activator of the enzyme, whereas GTP at low (less than 0.25 mM) concentrations increased the enzyme activity. The activation effect was negligible at higher concentrations of GTP. The calculated value of K0.5 of approx. 3 mM for unactivated enzyme decrased to approx. 0.6 mM and 1.1 mM when 0.5 mM ATP or 1.5 mM ADP were present in the incubation mixture, respectively. The theoretical model (Monod, J., Wyman, J. and Changeux, J.P. (1965) J. Mol. Biol. 12, 88-118) gave a partial explanation of these results.
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PMID:Regulatory properties of rat heart AMP deaminase. 44 47

The protein synthesis initiation factor eIF-3 (a multicomponent protein complex) was labelled with 32P by phosphorylation with a protein kinase present in a partially purified 'hemin-controlled repressor' preparation. The interaction of the labelled factor with the 40 S ribosomal subunit during the course of initiation was followed. It binds to the 40 S subunit in the absence of other initiation factors and inhibits the Mg2+-dependent reassociation of the 40 S with the 60 S ribosomal subunit. It stimulates the binding of the ternary complex (eIF-2, GTP, Met-tRNAf) to the 40 S subunit, and earlier work (Trachsel, H., Schreier, M.H., Erni, B. and Staehelin, T. (1977) J. Mol. Biol. 116, 745-767) also showed it to be essential for the subsequent binding of mRNA. The factor is released from the 40 S initiation complex during the 60 S subunit joining reaction.
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PMID:Initiation of mammalian protein synthesis. The multiple functions of the initiation factor eIF-3. 51 82

The paper summarizes studies of the molecular mechanism of the dynamic function of the ribosome, i. e. translocation, performed in the author's laboratory during the past decade. The hypothesis of the locking-unlocking of the ribosomal subparticles and the kinematical model of the working ribosome, the processes of spontaneous (factor-free) and factor-dependent translocation, the sequence of events in the factor-dependent translocation, the energetics of translocation and the contribution of the elongation factors with GTP are considered. The following conclusions are made: (1) the translocation mechanism is intrinsic to the structural organization of the ribosome itself but not introduced by the protein elongation factors; (2) the transpeptidation reaction is one of the sources of energy for the work of the translocation mechanism; (3) the protein elongation factors with GTP impart additional energy to the ribosome, including that for translocation, and thus ensure excess power which is realized, in particular, in the increase of the translocation rate and its resistance against inhibitors and hindrances; (4) the promoting role of the elongation factors with GTP does not proceed by a direct conjugation of GTP hydrolysis with translocation, but through the affinity of the elongation factors to the ribosome, with a subsequent compensation of the affinity at the expense of GTP cleavage.
Mol Biol (Mosk)
PMID:[Translocation mechanism of ribosomes]. 61 48

Rat liver nuclear 30 S ribonucleoprotein particles containing pre-mRNA and nuclear sap proteins have been shown to modify in vitro the synthetic dinucleotide ppGpC in the presence of GTP and S-adenosyl-L-methionine (SAM) by the formation of a blocked and methylated (capped) structure 7(meG(5')ppp(5'-GmepC. In the absence of SAM the predominant reaction was GpppGpC. Our results indicate that the 30S ribonucleoprotein particles (informofers) as well as the proteins of the nuclear sap possess both guanylyltransferase, N7-, and 2-o-methyltransferase activities.
Mol Biol Rep 1978 Jun 16
PMID:Methylated cap formation by enzymes bound to nuclear informofer particles. 68 86

We estimate that E. coli RNA polymerase is able to form stable, rifampicin-resistant, pre-intiation complexes with Adenovirus 2 DNA at three to six binding sites. The number of RNA chains initiated from such complexes has been determined form the incorporation of gamma-32P-ATP and -GTP at two rifampicin concentrations (7 mug/ml and 24 mug/ml) and after pre-incubation at either 25 or 37 degrees C. The total number of RNA chains initiated ranges from 2.6 per Ad 2 DNA molecule at a rifampicin concentration of 24 mug/ml and pre-incubation temperature of 25 degrees C, to 5.4 per Ad 2 DNA molecule at a rifampicin concentration of 7 mug/ml and pre-incubation temperature of 37 degrees C. Efficient initiation with GTP occurs only after pre-incubation at 37 degrees C whereas initiation with ATP is equally as efficient at either pre-incubation temperature. Promoters for initiation with ATP have been localized to the leftmost 58% of the Ad 2 DNA molecule, defined by the EcoR.RI restriction endonuclease fragment A; promoters for initiation with GTP are located on the remaining 42% of the Ad2 DNA molecule. It is likely that on Adenovirus 2 DNA each RNA chain is initiated from a unique binding site which constitutes a seperate promoter for E. coli RNA polymerase.
Mol Gen Genet 1976 Jan 16
PMID:In vitro transcription of adenovirus 2 DNA. II. Quantification and localization of promoters for E. coli RNA polymerase. 76 52

It is shown that ribosomes, the 30S subparticles of which are reconstituted without protein S12, read out poly(U) and synthesize polyphenylalanine in the absence of protein elongation factors (EF-T and EF-G) and GTP, i.e. perform "non-enzymatic" translation. On the contrary, ribosomes, the 30S subparticles of which are reconstituted with protein S12, do not display "non-enzymatic" translation without its activation with parachloromercuribenzoate. This means that a complete removal of protein S12 from the ribosome, as well as its damage with para-chloromercuribenzoate, leads to the unblocking of the potential ability of ribosomes for spontaneous ("non-enzymatic") translocation. The presence of intact protein S12 in the ribosome prevents spontaneous (EF-G-GTP-independent) translocation. A suggestion is made that the intact protein S12 forms an additional contact between the ribosomal subparticles and thus participates in the ribosomal mechanism of translocation by affecting the locking-unlocking of the subparticles.
Mol Biol (Mosk)
PMID:Studies on the mechanism of translocation in ribosomes. IV. The role of ribisomal proteins S12 in translocation. 76 74

The paper deals with the comparative investigation of initiation and in vitro RNA synthesis on DNA template by E. coli RNA polymerase and B-form of calf thymus RNA polymerase. It was shown in hybridization experiments that in the range of Cot values between 10(2) and 10(4) RNA synthesized by calf thymus RNA polymerase was hybridized with homologous DNA more effectively than RNA synthesized by E. coli RNA polymerase. No differences were observed in the case of low Cot values. RNA chains synthesized by calf thymus RNA polymerase contained in average about 300-600 nucleotides per chain as determined in the kinetic experiments with ATP-gamma-32P and GTP-gamma-32P. These values are in average 5-10 times lower than in the case of bacterial enzyme. The data presented show that calf thymus and E. coli RNA polymerases initiate the RNA synthesis at apparently different sites of calf thymus DNA. The results obtained make the possibility of specific transcription of eucaryotic DNA by bacterial RNA polymerase to be doubtful.
Mol Biol (Mosk)
PMID:[Transcription of DNA by RNA polymerases of E. coli and calf thymus]. 76 44

Treatment of elongation factor G (EF-G) with the thiol reagent N-ethylmaleimide only partially inhibits (10 to 70%) the activity of the factor in (a) guanosine nucleotide-EF-G-ribosome complex formation, (b) uncoupled ribosome-dependent GTP hydrolysis, and (c) polypeptide synthesis. Moreover, a similar treatment of the factor with N-[3H]ethylmaleimide does not lead to 3H-label being associated with a GDP-EF-G-ribosome-fusidic acid complex. Thus, the results indicate the presence in EF-G preparations of a form of the factor that does not react with N-ethylmaleimide.
Mol Biol Rep 1976 Apr
PMID:A form of elongation factor G insensitive to N-ethyl-maleimide. 77 17

A protein kinase specific for casein and acidic ribosomal proteins was isolated and partly characterized. It was found that the enzyme utilizes GTP and ATP as phosphoryl donors. Its affinity for ATP was considerably higher than for GTP with the km values of 7.6 X 10(-6)M and 5.5 X 10(-5)M, respectively. Two-dimensional acrylamide gel electrophoresis revealed the phosphorylation of the same ribosomal proteins with either of the [gamma-32P] nucleotides used. It was also shown that one acidic protein (S1 or S2) of 40 S and two acidic proteins (L2 and L3) of 60 S ribosomal subunits were predominantly phosphorylated in vitro. The phosphorylated proteins: L2 and L3 seem to correspond to the proteins of L7 and L12 of E. coli ribosomes. The isolated kinase phosphorylated several basic ribosomal proteins though to a lower extent than the acidic ones.
Mol Biol Rep 1976 Nov
PMID:Ribosomal protein as substrate for a GTP-dependent protein kinase from yeast. 79 85


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