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J Chroboczek

Publications and source records attributed to J Chroboczek.

52 records · Page 3Linked to original sources
PubMed1988

Determination of the nucleotide sequence for the penton-base gene of human adenovirus type 5.

The major structural proteins of adenovirus (Ad), which form the external capsid, are hexon, penton base and fiber. The primary structure of the Ad5 penton base has been deduced from the nucleotide sequence of the corresponding gene. It has 98.6% homology with the sequence of the analogous protein from Ad2. This result is in contrast with the significantly lower homology found for the two other major structural proteins, the hexon and the fiber.

Adenoviruses, Human↗
PubMed1987

The sequence of adenovirus fiber: similarities and differences between serotypes 2 and 5.

The amino acid sequence of the fiber from adenovirus type 5 has been deduced from the nucleotide sequence of the fiber gene. This sequence is compared with that of adenovirus type 2, a closely related serotype. We find 69% homology for the fiber protein whereas the known nonstructural proteins of these two serotypes have 99% sequence homology. A detailed sequence analysis was performed in the context of the model proposed by Green et al., [(1983), EMBO J., 8, 1357-1365] for the folding of the polypeptide chain of the adenovirus type 2 fiber. The N-terminal region, which in the virion is associated with the capsid, is identical for these two serotypes. In the shaft of the fiber the features, such as periodicity of the prolines and of the hydrophobic residues in the amino acid sequence, on which the model for the adenovirus type 2 is based are very well preserved in adenovirus type 5. On the other hand, there are large differences all along the sequence of the shaft of the fiber showing that there is a very limited homology between the amino acids in the two serotypes when they do not have a key role in establishing the structure. In the knob the homology between serotypes is 64%. These results are consistent with the differences between serotypes being confined to the exposed proteins.

Adenoviruses, Human↗
PubMed1986

Identification of regions of brome mosaic virus coat protein chemically cross-linked in situ to viral RNA.

RNA-protein cross-links were introduced into brome mosaic virus in situ by using the heterobifunctional agent p-azidophenylglyoxal. An improved RNA isolation method, without phenol extraction, was used to isolate RNA cross-linked with protein. RNA of the covalently linked complex was acid-digested and the oligonucleotides still attached to protein were 5'-end-labelled with 32P. The complexes were digested with trypsin and the tryptic peptides were purified by reversed-phase high-performance liquid chromatography. Amino acid analyses of cross-linked tryptic peptides revealed that out of the total 188 amino acids of brome mosaic virus coat protein only the 80 N-terminal amino acids are involved in the interaction with viral RNA. These results are discussed in connection with a predicted secondary structure of the coat protein. Both alpha helix (for amino acids 11-19) and other structures (between amino acids 20 and 80) are implicated in the coat protein-viral RNA interactions.

Amino Acids↗
PubMed1986

Human adenovirus 2 temperature-sensitive mutant 112 contains three mutations in the protein IIIa gene.

The temperature-sensitive (ts) mutant 112 of human adenovirus 2 is defective in the late stage of virus maturation. The region of functional mutation has been localised by marker rescue. It was observed that the ts mutation can be rescued by the left-hand part of the wild-type gene (nucleotides 12,301-12,891). By nucleotide sequencing, two mutations, both C to T (at position 12,386 and 12,741), were found in this region. The first one, in the glycine 20 codon, is silent, whereas the second changes alanine 145 to valine. A third mutation, which changed C to A (nucleotide 13,613), was identified in the right-hand part of the gene, resulting in the replacement of alanine-436 by threonine.

Adenoviruses, Human↗
PubMed1985

Incomplete polypeptides are formed in vitro by premature chain termination.

The mechanism of incomplete polypeptides formation during protein synthesis was studied in the wheat germ cell-free system programmed with brome mosaic virus RNA 4. The synthesis of coat protein, the complete product of RNA 4 translation, was accompanied by the appearance of polypeptides of lower molecular mass. It was shown that incomplete products are formed by translation of different lengths of RNA 4, always from the first 5' AUG codon, and were due neither to proteolysis of coat protein nor to the translation of nucleolytic fragments of mRNA. The molecular masses of incomplete products were determined and the nucleotide sequence of RNA 4 was examined in the regions where wheat germ ribosomes stop translating. It was found that they contained, on average, a slightly higher guanosine content than the total coding part of RNA 4. Translation of RNA 4 in the reticulocyte lysate resulted in a marked diminution of incomplete polypeptides. Addition of high-speed supernatant from reticulocyte lysate prevented the formation of incomplete products during translation of RNA 4 in the wheat germ system. This suggests that reticulocyte lysate contains some factor(s) which facilitate the movement of ribosomes beyond the regions where the elongation is retarded.

Animals↗
PubMed1985

Effect of dinucleotides on wheat germ translation system.

The effect of ribodinucleoside monophosphates on total protein synthesis was studied in a wheat germ cell-free system, using brome mosaic virus (BMV) RNA as a messenger. Dinucleotides inhibit total protein synthesis to different extents. Of those tested the most inhibitory is CpA. The inhibitory effect of dinucleotides is due to their adverse effect on initiation and not on elongation of polypeptide synthesis. It seems that the dinucleotides complementary to the initiation codon are able to compete with the initiator tRNA during initiation of protein synthesis. The comparison of the effect exerted by different dinucleotides suggests that under conditions of the in vitro protein synthesis RNA 4 is an mRNA molecule with the initiation codon and its immediate neighbourhood being exposed.

Cell-Free System↗
PubMed1982

Effect of secondary structure of messenger ribonucleic acid on the formation of initiation complexes with prokaryotic and eukaryotic ribosomes.

The effect of modification of the secondary structure of phage f2 RNA and brome mosaic virus (BMV) RNA 3 on the formation of initiation complexes in Escherichia coli and wheat germ protein-synthesizing systems was studied. Modification of the RNAs was achieved by using O-methylhydroxylamine, which specifically reacts with cytosines; this leaves the initiation codons unchanged and, under denaturing conditions, leads to irreversible unfolding of the RNA. E. coli ribosomes interact with newly exposed AUG/GUG codons in the modified templates forming polysomes, whereas they form monosomes with native f2 RNA or BMV RNA 3. With wheat germ ribosomes, disomes are formed in the presence of BMV RNA 3, either native or modified. With f2 RNA, eukaryotic ribosomes form monosomes, independent of the secondary structure of the template. The results indicate that, in contrast to prokaryotic ribosomes, binding of eukaryotic ribosomes to f2 RNA or BMV RNA 3 is not affected by modification of the secondary structure of these messengers.

Cells↗
PubMed1980

Regulation of brome mosaic virus gene expression by restriction of initiation of protein synthesis.

The translation of total and individual brome mosaic virus (BMV) RNAs was examined in a wheat germ cell-free system in the presence of various inhibitors. Inhibitors of the initiation of polypeptide synthesis, e.g., potassium ions, 7-methylguanosine 5' -monophosphate, and aurintricarboxylic acid, were shown not only to inhibit overall BMV protein synthesis but also to change the ratio of BMV polypeptides synthesized. Under conditions restrictive for initiation, the translation of nonstructural BMV genes was suppressed, but coat protein synthesis proceeded at a high rate. A similar discrimination among BMV messengers was exerted by a regulatory protein kinase isolated from wheat germ. These results suggest that the regulation of the expression of BMV genes is based on a difference in the mechanism of formation of initiation complexes for individual BMV messages.

Journal Article↗
PubMed1978

The host cell fraction that increases the infectivity of bacteriophage f2.

A fraction that increases infectivity of bacteriophage f2 was isolated from uninfected E. coli cells. The greatest effect was obtained when the fraction was added to the phage reconstituted in vitro. The fraction isolated from the ribosome-free supernatant consisted of proteins, lipids, carbohydrates, and unidentified material. Cleavage of protein by the treatment with trypsin did not significantly affect the infectivity-restoring activity. It is suggest that lipids may play an essential role in the activity of the fraction isolated from the host cell.

Bacteriophages↗
PubMed1977

Effect of rifampicin on the infectivity of RNA bacteriophage f2.

RNA bacteriophage f2, treated in vitro with rifampicin, loses infectivity dramatically. Rifampicin interacts with phage RNA, binding to a few specific sites. Inhibition of phage RNA infectivity occurs at 10-100 times lower molar excess of rifampicin than inhibition of infectivity of intact phage particles. Thus the phage capsid acts as a barrier, diminishing interaction of the drug with phage RNA.

Coliphages↗
PubMed1975

Template activity of complexes formed between bacteriophage f2 RNA and coat protein.

Formation of complexes between f2 RNA polymerase cistron was partially inhibited, some RNA and coat protein was studied using salt conditions which are optimum for phage protein synthesis. In this ionic environment, coat protein precipitation can be prevented by sulfhydryl group-protecting agents. Complexes formed at different protein-RNA input molar ratios were isolated and tested for template activity in an in vitro protein synthesizing system. Simultaneously, the number of protein molecules bound per RNA strand in such complexes was measured by the membrane (Millipore) filtration technique. Under conditions in which translation of the RNA strands were complexed with six molecules of coat protein, whereas some remained unbound. Strong inhibition of the translation of the RNA polymerase cistron was observed when each of the RNA strands present in the mixture was associated with six molecules of coat protein.

Carbon Radioisotopes↗
PubMed1975

Hexamer of bacteriophage f2 coat protein as a repressor of bacteriophage RNA polymerase synthesis.

Formation of complex I between phage f2 RNA and coat protein, leading to repression of phage RNA polymerase synthesis, depends nonlinearly upon the concentration of the coat protein. Maximum formation of complex I was observed when six molecules of coat protein were bound to one molecule of RNA. RNase digestion of a glutaraldehyde-fixed complex left, as the products, coat protein oligomers. The heaviest, hexamers, predominated in the mixture. It was also shown that, in an ionic environment required for phage protein synthesis, coat protein at a concentration optimum for complex I formation exists in solution as a dimer. The results indicate that the translational repression of the RNA polymerase cistron is due to a cooperative attachment to phage template of three dimers of coat protein, forming a hexameric cluster on an RNA strand.

Centrifugation, Density Gradient↗
PubMed1973

Specificity of formation of complexes between coat protein and bacteriophage f2 RNA.

The specificity of formation of phage f2 RNA-protein complexes was studied. Complex I contains up to 8 mol of coat protein per 1 mol of RNA. Its formation proceeds equally well in medium (i) without magnesium ions, (ii) containing magnesium ions, (iii) containing 4 mM EDTA, and (iv) at temperatures from 0 to 45 C. Complex II contains up to 200 mol of coat protein per 1 mol of RNA. Its formation is inhibited by the presence of magnesium ions in medium. Formaldehyde- or methoxyamine-treated f2 RNA in which only exposed bases were modified showed a normal pattern of complex II formation, whereas formation of complex I was inhibited or abolished. We conclude that complex I formation involves the interaction between coat protein and specific region of exposed bases in RNA. A possible site of attachment of coat protein is discussed.

Binding Sites↗
PubMed1973

[Rifampin].

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Animals↗