PubMed Health⌕ Search

Biomedical subjects

T Twardowski

Publications and source records attributed to T Twardowski.

At least 19 recordsLinked to original sources
PubMed2001

Stress conditions applied to the interpretation of translation machinery.

Gene expression is regulated at the critical steps: a regulatory event occurs at the step which has a critical effect and is responsible for the limiting rate. Enzyme activity can be regulated at several different levels: transcriptional, translational or post-translational. In this review we describe (and illustrate with experimental data) plant stress which induces regulatory mechanisms at the translational and post-translational levels. We found evidence for autorepression regulatory system of ferritin biosynthesis. Based on the knowledge of the molecular mechanism of regulation, we believe that ferritin protects the environment against heavy metal ions and supplements biological system(s) with iron. The quinolizidine alkaloids' (QA) biosynthesis is lysine decarboxylase (LDC)-dependent. The available pool of LDC limits the conversion of lysine to cadaverine. The amount of LDC depends on transcriptional and translational efficiency. However, in the light of the presented data, we have evidence for a post-translational regulatory system, i.e. the activation of LDC from low to high activity enzyme through the conversion from higher to lower molecular weight form. The plant protection system is very efficient. Understanding of the defence systems such as plant response to stress, should provide us with a possibility of applying this knowledge in practice and finding novel applications.

Alkaloids↗
PubMed2001

[Cloning - controversies].

Cloning of the human being is not only highly controversial; in the opinion of the authors it is impossible - we are not able to reproduce human behaviour and character traits. Reproduction through cloning is limited to personal genome resources. The more important is protection of genomic characteristics as private property and taking advantage of cloning for production of the human organs directly or through xenotransplants. In this paper we present the legislation related to cloning in Poland, in the European Union and other countries. We also indicate who and why is interested in cloning.

Bioethical Issues↗
PubMed2000

Design of peptides with high affinities for heparin and endothelial cell proteoglycans.

Proteoglycan-binding peptides were designed based on consensus sequences in heparin-binding proteins: XBBXBX and XBBBXXBX, where X and B are hydropathic and basic residues, respectively. Initial peptide constructs included (AKKARA)(n) and (ARKKAAKA)(n) (n = 1-6). Affinity coelectrophoresis revealed that low M(r) peptides (600-1,300) had no affinities for low M(r) heparin, but higher M(r) peptides (2,000-3,500) exhibited significant affinities (K(d) congruent with 50-150 nM), which increased with peptide M(r). Affinity was strongest when sequence arrays were contiguous and alanines and arginines occupied hydropathic and basic positions, but inclusion of prolines was disruptive. A peptide including a single consensus sequence of the serglycin proteoglycan core protein bound heparin strongly (K(d) congruent with 200 nM), likely owing to dimerization through cysteine-cysteine linkages. Circular dichroism showed that high affinity heparin-binding peptides converted from a charged coil to an alpha-helix upon heparin addition, whereas weak heparin-binding peptides did not. Higher M(r) peptides exhibited high affinities for total endothelial cell proteoglycans (K(d) congruent with 300 nM), and approximately 4-fold weaker affinities for their free glycosaminoglycan chains. Thus, peptides including concatamers of heparin-binding consensus sequences may exhibit strong affinities for heparin and proteoglycans. Such peptides may be applicable in promoting cell-substratum adhesion or in the design of drugs targeted to proteoglycan-containing cell surfaces and extracellular matrices.

Amino Acid Sequence↗
PubMed1998

tRNA aminoacylated at high pressure is a correct substrate for protein biosynthesis.

tRNA can be aminoacylated specifically with amino acids at high pressure of 6 kbar (1 bar = 1.013 atm = 0.1 MPa = 10(5) Pa) in the absence of the specific aminoacyl-tRNA synthetase and ATP. In this paper we present new evidence obtained by HPLC chromatography and TLC analysis that the esterification reaction under pressure really takes place at the 3' end of the tRNA molecule. If so, tRNA to be aminoacylated undergoes conformational changes similar to those induced with aminoacyl-tRNA synthetase. The most important finding is that aminoacyl-tRNA obtained at high pressure binds to ribosomes and participates in the synthesis of polyphenylalanine in vitro. This is the best proof of proper charging of tRNA at high pressure.

Acylation↗
PubMed1998

Functional role of rRNAs in plant translation system tested with antisense strategy.

There are regions in rRNA which are evolutionary conserved and exposed on ribosomal surface. We selected in plant material (Lupinus luteus) two of them: the alpha-sarcin domain of 26S rRNA (L-rRNA) and C loop of 5S rRNA, to be further investigated using antisense oligomers as research tools. We found inhibition of the model polypeptide biosynthesis (up to 80%) due to specific hybridization of oligomers addressed to alpha-sarcin domain and loop C. Based on our results we present the evidence for the key role played by these regions of rRNAs during protein biosynthesis in plant system. According to our hypothesis, conformational changes of these two regions are synchronised and cooperative during transition of pre- to post-translocation state of the ribosome. The correlation of structure and activity of rRNA domains in translation is shown.

Base Sequence↗
PubMed1996

Effects of antisense DNA against the alpha-sarcin stem-loop structure of the ribosomal 23S rRNA.

Antisense DNAs complementary against various sequences of the alpha-sarcin domain (C2646-G2674) of 23S rRNA from Escherichia coli were hybridized to naked 23S rRNA as well as to 70S ribosomes. Saturation levels of up to 0.4 per 70S ribosome were found, the identical fraction was susceptible to the attack of the RNase alpha-sarcin. The hybridization was specific as demonstrated with RNase H digestion, sequencing the resulting fragments and blockage of the action of alpha-sarcin. The RNase alpha-sarcin seems to approach its cleavage site from the 3' half of the loop of the alpha-sarcin domain. Hybridization is efficiently achieved at 37 degrees C and can extend at least into the 3' strand of the stem of the alpha-sarcin domain. However, the inhibition of alpha-sarcin activity is observed at 30 degrees C but not at 37 degrees C. For a significant inhibition of poly(Phe) synthesis the temperature had to be lowered to 25 degrees C. The results imply that the alpha-sarcin domain changes its conformation during protein synthesis and that the conformational changes may include a melting of the stem of the alpha-sarcin domain.

Base Sequence↗
PubMed1994

Evidence for plant ribosomal 5S RNA involvement in elongation of polypeptide chain biosynthesis.

A series of short oligo-DNA probes (8-10-mers) complementary to various regions of the plant ribosomal 5S ribonucleic acid (5S rRNA) have been synthesized. The results of their hybridization to free 5S rRNA and to ribosomes pointed to the availability of nucleotides in loop "C" for complexation. We found a correlation between hybridization of selected oligonucleotides and their inhibitory effect on enzymatic binding of Phe-tRNA and poly(Phe) synthesis on wheat germ 80S ribosomes. Evidence was obtained for involvement of 5S rRNA in the elongation of polypeptide chain during protein biosynthesis. 5S rRNA seems to play a critical role in protein biosynthesis, probably through causing conformational changes of loop C.

Antisense Elements (Genetics)↗
PubMed1992

The two main states of the elongating ribosome and the role of the alpha-sarcin stem-loop structure of 23S RNA.

According to the allosteric three-site model of the elongation cycle the ribosome oscillates between two main-functional states, viz the pre-translocational state with occupied A and P sites (E site with low affinity) and the post-translocational state with occupied P and E sites (A site with low affinity). This proposition could be confirmed by a determination of the thermodynamic parameters. High activation-energy barriers were found between both states, namely about 90 kJ mol-1 at 15 mM Mg2+ for either transition (post----pre transition = A-site binding and pre----post transition = translocation). The various A-site states (binding of ternary complex, EF-Tu dependent GTP cleavage, peptide-bond formation) are not separated by significant activation-energy barriers. The rate-limiting step of the elongation cycle is A-site binding, and not translocation as assumed previously. The principal role of both elongation factors is the reduction of the respective activation-energy barrier, thus accelerating the rate of the elongation cycle by several orders of magnitude. Cleavage of a single phosphodiester bond after G2661 of 23S rRNA by the RNase alpha-sarcin abolishes the functions of both elongation factors on the ribosome. This observation implies that the alpha-sarcin stem-loop structure plays an important role in the ribosomal conformational changes involved in the allosteric transitions. Indeed we could demonstrate that suitable oligodeoxynucleotide probes complementary to the alpha-sarcin region induce a conformational change in the 50S subunits; this conformational change causes an irreversible dissociation of tightly coupled ribosomes upon sucrose-gradient centrifugation.(ABSTRACT TRUNCATED AT 250 WORDS)

Allosteric Site↗
PubMed1988

Immunochemical properties of elongation factors 1 of plant origin.

Elongation factors 1 (EF-1) have been isolated from different plants: wheat, yellow lupine, blue lupine, Chinese cabbage and Norway maple. Antibodies for EF-1 from yellow lupine have been obtained in rabbits; antibodies for wheat EF-1 were elicited in mice. The immunological properties of EF-1 were assayed by the following methods: western blotting, double immunodiffusion and rocket immunoelectrophoresis. Our results suggest that one antigenic site is similar for all plant elongation binding factors tested. This epitope probably overlaps the centre of biological activity of EF-1, as was shown for wheat EF-1. The hypothesis concerning the potential presence of plant EF-1 as a subunit of turnip yellow mosaic virus RNA replicase (similar to prokaryotic EF-Tu in the Q beta RNA replicase system) has also been tested using immunotechniques as well as tests of biological activity, but has not been confirmed.

Cross Reactions↗
PubMed1988

The inhibitory effect of oligodeoxynucleotides complementary to different fragments of tRNA structure on the enzymatic binding of Phe-tRNA to poly-U-programmed eukaryotic ribosomes.

The effect of several oligodeoxynucleotides complementary to the fragments of yellow lupin tRNA(Phe) was tested in the aminoacylation of tRNA(Phe) and in the binding of Phe-tRNA(Phe) to poly-U-programmed eukaryotic ribosomes. Oligonucleotides tested in the aminoacylation test did not give any inhibition. Monomers and dimers did not have any significant influence on the binding assay, either. A different percentage of inhibition of the binding of Phe-tRNA to ribosomes has been observed for oligonucleotides. Heptamer complementary to the anticodon loop gave 100% inhibition of the binding reaction. However, the oligonucleotides complementary to both the anticodon loop and stem and longer than the heptamer were much less effective inhibitors. A high inhibitory effect was also observed for trimers and for the decamer complementary to the D-loop and CCA-end.

Base Sequence↗