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C B Bruni

Publications and source records attributed to C B Bruni.

At least 91 records · Page 5Linked to original sources
PubMed1986

Gene structure in the histidine operon of Escherichia coli. Identification and nucleotide sequence of the hisB gene.

The bifunctional enzyme imidazoleglycerolphosphate dehydratase and histidinolphosphate phosphatase is encoded by the hisB gene. The fourth gene of the histidine operon, hisB, was cloned and mapped on a 2,300 base pair DNA fragment. In the present study we report the complete nucleotide sequence of the hisB gene of Escherichia coli. The gene is 1,068 nucleotides long and codes for a protein of 355 amino acids with an apparent molecular weight of 39,998 daltons. The protein product(s) of the hisB region of both Salmonella typhimurium and E. coli were identified by subcloning and expression in an in vitro translation system. In both organisms the hisB gene directed the synthesis of a single protein with an apparent molecular weight of 40,500 daltons, consistent with the data derived from the nucleotide sequence analysis.

Amino Acid Sequence↗
PubMed1986

Nucleotide sequence of the Escherichia coli hisD gene and of the Escherichia coli and Salmonella typhimurium hisIE region.

In this paper we report the nucleotide sequence of the hisD gene of Escherichia coli and of the his IE region of both E. coli and Salmonella typhimurium. The hisD gene codes for a bifunctional enzyme, L-histidinol:NAD+ oxidoreductase, of 434 amino acids with a molecular mass of 46,199 daltons. We established that the hisIE region of both S. typhimurium and E. coli is composed of a single gene and not, as previously believed, of two separate genes. The derived amino acid sequence indicates that the hisIE gene codes for a bifunctional protein of 203 amino acids with an approximate molecular mass of 22,700 daltons. We also determined the nucleotide sequence of a deletion mutant in S. typhimurium which abolishes the hisF and hisI functions but retains the hisE function. We deduced that the mutant produces a chimeric protein fusing the aminoterminal region of the upstream hisF gene to the carboxyl-terminal domain of the hisIE gene which encodes for the hisE function. In view of these results the structural and functional organization of the histidine operon in enteric bacteria needs to be revised. The operon is composed of only 8 genes and the pathway leading to the biosynthesis of the amino acid requires 11 enzymatic steps.

Amino Acid Sequence↗
PubMed1986

Primary and secondary structural homologies between the HIS4 gene product of Saccharomyces cerevisiae and the hisIE and hisD gene products of Escherichia coli and Salmonella typhimurium.

A detailed comparative analysis of the Escherichia coli and Salmonella typhimurium hisIE and hisD gene products and the functionally equivalent, single, HIS4 gene product of Saccharomyces cerevisiae permitted several insights concerning the relationship between these genes. Our analysis supports the idea that HIS4 results from the fusion of his IE and hisD. The comparison permitted a more precise definition of the functional domains of hisI/HIS4A and hisE/HIS4B as well as the two functional domains of hisD/HIS4C. The homologies between the bacterial and yeast sequences suggest a region of the hisD/HIS4C protein that may constitute one of the active centres. A large fragment at the amino terminal region of the yeast protein is missing from the bacterial hisIE gene product and is probably not needed for catalytic activity. Another region of non-homology in the yeast protein is probably a peptide bridge connecting the HIS4AB domain to HIS4C. Although the overall homology at the level of amino acid sequence is modest (about 38%) there is a striking similarity when the hydropathic patterns and predicted secondary structural configurations of these proteins are compared.

Amino Acid Sequence↗
PubMed1986

Coordinate developmental regulation of high and low molecular weight mRNAs for rat insulin-like growth factor II.

Insulin-like growth factor II (IGF-II) is a mitogenic polypeptide that is thought to play a role in fetal growth and development. To study the hormonal and developmental regulation of IGF-II gene expression, we have isolated a cDNA clone for rat IGF-II (rIGF-II) from a 12S [1.2-kilobase-pair (kbp)] fraction of mRNA from a rat liver cell line (BRL-3A) that directs the cell-free synthesis of pre-pro-rIGF-II. In the present study, the rIGF-II probe was used to determine the size of IGF-II RNA. Surprisingly, in BRL-3A cells and in neonatal liver, the probe hybridized under stringent conditions 10-20 times more strongly to a larger (4 kbp) RNA than to 1.2-kbp RNA. The 4-kbp RNA is almost exclusively cytoplasmic and is colinear with a 551-base fragment of the rIGF-II cDNA insert containing coding and 3' noncoding regions. The 4-kbp and 1.2-kbp RNA species are regulated coordinately with developmental age, being high in liver from neonatal rats but not detectable in liver from older animals, suggesting that both IGF-II mRNA species arise from a single primary transcript by alternative RNA processing. Although oligodeoxynucleotide hybridization and S1 nuclease protection experiments suggest that the 4-kbp RNA contains an intact protein-coding region, fractions enriched in 4-kbp RNA do not direct the translation of pre-pro-rIGF-II in vitro. This may indicate that the 4-kbp RNA specifies an altered protein product that has not yet been recognized, or alternatively that it contains a normal protein-coding region but requires further RNA processing to be activated for translation.

Animals↗
PubMed1985

Convergently functional, Rho-independent terminator in Salmonella typhimurium.

A typical Rho-independent terminator of transcription was found at the end of the histidine operon of Salmonella typhimurium. This site is used to terminate, in addition to the his operon mRNA, a 1,200-nucleotide RNA of unknown function transcribed on the opposite strand. The efficiency of termination of transcription at this site was investigated in vivo and in vitro by cloning of the terminator structure in either orientation in vector systems used to study regulatory signals. Termination of transcription at this site was very efficient, both in vivo and in vitro, and in both orientations.

Chromosome Mapping↗
PubMed1985

Regulation of single and multicopy his operons in Escherichia coli.

We fused segments of the Escherichia coli his regulatory region to galK in single-copy and multicopy vectors. These fusions demonstrated that (i) derepression of his by histidine starvation is due exclusively to attenuation; (ii) the his promoter is metabolically regulated; and (iii) both regulatory systems operate when the his regulatory region is present on a multicopy plasmid. Thus, there is no evidence for titration of his regulatory elements. Deletions of the his anti-attenuator region, carried on multicopy plasmids, cause low-level galK expression. This expression is not stimulated by histidine starvation, but is growth rate dependent. We replaced the his attenuator with the efficient lambda terminator, to. In the context of the his regulatory region, however, lambda to only partially terminates transcription.

Bacteriophage lambda↗
PubMed1985

Cloning, structure, and expression of the Escherichia coli K-12 hisC gene.

We used an expression vector plasmid containing the Escherichia coli K-12 histidine operon regulatory region to subclone the E. coli hisC gene. Analysis of plasmid-coded proteins showed that hisC was expressed in minicells. A protein with an apparent molecular weight of 38,500 was identified as the primary product of the hisC gene. Expression was under control of the hisGp promoter and resulted in very efficient synthesis (over 100-fold above the wild-type levels) of imidazolylacetolphosphate:L-glutamate aminotransferase, the hisC gene product. The complete nucleotide sequence of the hisC gene has been determined. The gene is 1,071 nucleotides long and codes for a protein of 356 amino acids with only one histidine residue.

Amino Acid Sequence↗
PubMed1983

Gene organization in the distal part of the Salmonella typhimurium histidine operon and determination and sequence of the operon transcription terminator.

Several transducing phages, carrying different deletions of the Salmonella typhimurium histidine operon were constructed and mapped. These phages were used to obtain fragments of DNA comprising different regions of the operon, which were subcloned in plasmid vectors. The recombinant plasmids allowed the construction of a physical and restriction map of the histidine operon. The presence of the different genes on individual fragments was confirmed by complementation tests. The transcription termination site of the histidine operon has been established by S1 mapping and sequence analysis. The entire operon measures about 7100 base pairs and the last six structural genes are contained in 3450 bases of genetic materials.

Chromosome Mapping↗
PubMed1983

Structure and function of the internal promoter (hisBp) of the Escherichia coli K-12 histidine operon.

The entire histidine operon of Escherichia coli K-12 was cloned in the vector plasmid pBR313, and a complete restriction map of the operon was determined. By using subclones, complementation tests, and enzyme assays, we were able to make a correlation between the physical map and the genetic map of the operon. We determined the sequence of a fragment of DNA 665 base pairs long, comprising the distal portion of the hisC gene, the proximal portion of the hisB gene, and the internal transcription initiation site hisBp. The efficiency of this promoter was assessed under different physiological conditions by cloning the DNA fragment in a recombinant vector system used to study transcriptional regulatory signals. The precise point at which transcription initiates was determined by S1 nuclease mapping.

Base Sequence↗
PubMed1982

Cloning and expression of the distal portion of the histidine operon of Escherichia coli K-12.

The operator-distal genes hisBHAFI(E) of the Escherichia coli K-12 histidine operon were mapped on a DNA fragment 4,500 base pairs long. This fragment, originally present in a lambda transducing phage, was cloned in the vector plasmid pBR313. A restriction map was determined, allowing identification of the orientation of the genes in the fragment. The cloned genes were expressed in appropriate hosts, independent of the orientation of the DNA fragment, as shown by transformation tests and by enzyme assays of one of the gene products, hisB, histidinol phosphatase. An internal transcription initiation site was identified by isolation of the cellular RNA, hybridization to specific DNA probes, and mapping by S1 nuclease.

Cloning, Molecular↗
PubMed1982

Cell-free synthesis of rat insulin-like growth factor II.

Total RNA extracted from a rat liver cell line (BRL 3A) that synthesizes rat insulin-like growth factor II (IGF-II) was translated in a reticulocyte lysate cell-free system. Incubation of the translation products with antisera to mature rat IGF-II (Mr 8700) selectively immunoprecipitated a Mr 21,600 protein. We propose that this protein represents pre-pro-IGF-II.

Animals↗
PubMed1981

Identification, nucleotide sequence and expression of the regulatory region of the histidine operon of Escherichia coli K-12.

A restriction fragment has been isolated and its nucleotide sequence determined. This fragment contains sites for RNA polymerase binding, initiation and termination of transcription of the Escherichia coli histidine operon. In vitro transcription of plasmids containing this region generates one single histidine-specific, attenuated, small RNA: the leader RNA. This RNA is more efficiently transcribed when the template DNA is supercoiled. Another promoter was identified on the same fragment of deoxyribonucleic acid by in vitro transcription, DNA sequencing and RNA polymerase binding. Both promoters, transcribing in opposite direction, are very A-T rich and are separated by a G-C rich region containing a palyndromic structure.

Base Sequence↗
PubMed1981

In vivo and in vitro detection of the leader RNA of the histidine operon of Escherichia coli K-12.

The DNA of the attenuator region of the histidine operon of Escherichia coli has been transcribed in a purified in vitro system and found to synthesize two major RNA transcripts. The first one, 180 nucleotides long, has been identified as the histidine-specific leader RNA. It contains the coding sequence for the leader peptide [Di Nocera, P. P., Blasi, F., Di Lauro, R., Frunzio, R. & Bruni, C. B. (1978) Proc. Natl. Acad. Sci. USA 75, 4276-4280] and is terminated at the attenuator site. Termination of transcription at this site is extremely efficient in the in vitro system. The leader RNA also has been detected in vivo in a minicell producer strain transformed with plasmids harboring the regulatory region of the histidine operon of E. coli. A second RNA molecule is synthesized in the in vitro system. It has a divergent direction of transcription with respect to the histidine leader RNA, but its role, if any, in the regulation of the histidine operon remains to be ascertained. The existence of the histidine leader RNA lends support to the regulatory mechanism which postulates that regulation of the histidine operon is dependent on the alternative secondary structures that the leader RNA may assume, depending on whether or not the histidine-rich leader peptide is translated.

Base Sequence↗
PubMed1980

Structural and physiological studies of the Escherichia coli histidine operon inserted into plasmid vectors.

A fragment of deoxyribonucleic acid 5,300 base paris long and containing the promoter-proximal portion of the histidine operon of Escherichia coli K-12, has been cloned in plasmid pBR313 (plasmids pCB2 and pCB3). Restriction mapping, partial nucleotide sequencing, and studies on functional expression in vivo and on protein synthesis in minicells have shown that the fragment contains the regulatory region of the operon, the hisG, hisD genes, and part of the hisC gene. Another plasmid (pCB5) contained the hisG gene and part of the hisD gene. Expression of the hisG gene in the latter plasmid was under control of the tetracycline promoter of the pBR313 plasmid. The in vivo expression of the two groups of plasmids described above, as well as their effect on the expression of the histidine genes not carried by the plasmids but present on the host chromosome, has been studied. The presence of multiple copies of pCB2 or pCB3, but not of pCB5, prevented derepression of the chromosomal histidine operon. Possible interpretations of this phenomenon are discussed.

Base Sequence↗
PubMed1979

Characterization of a somatomedin (insulin-like growth factor) synthesized by fetal rat liver organ cultures.

Explants of 19- to 20-day fetal rat liver synthesize polypeptides biochemically and immunologically related to the well characterized somatomedin (insulin-like growth factor) BRL-MSA, multiplication-stimulating activity. Fetal MSA was purified from media conditioned by fetal liver explants by chromatography on Sephadex G-75 under acid conditions. Partially purified fetal MSA: 1) inhibited the binding of BRL-MSA to the MSA receptor of rat liver plasma membranes, to somatomedin-binding proteins from rat serum, and to rabbit anti-BRL-MSA serum; 2) had a molecular weight of 4,500 to 12,500 determined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate; 3) stimulated the incorporation of [3H]thymidine into the DNA of chick embryo fibroblasts and induced cell multiplication; 4) stimulated glucose oxidation in rat adipocytes and weakly inhibited the binding of insulin to the insulin receptors of IM-9 lymphocytes; and 5) stimulated sulfate uptake in costal cartilage from hypophysectomized rats. These activities were associated with the same molecular species in fetal MSA preparations following disc acrylamide electrophoresis and co-migrated with active BRL-MSA peptides.

Animals↗
PubMed1978

Nucleotide sequence of the attenuator region of the histidine operon of Escherichia coli K-12.

The attenuator region of the histidine operon of Escherichia coli K-12 has a potential coding capacity for two peptides, one of 16 amino acids and another of 30 amino acids. This region is followed by a perfect palindrome of 14 base pairs separated by five nucleotides. A G+C-rich region precedes and follows a possible transcription termination sequence. These features are compatible with a model in which active translation of a leader mRNA interferes with transcription termination, thus causing derepression of the histidine operon. The sequence of the region coding for the hypothetical 16-amino acid peptide is of particular relevance because it indicates the site and a possible mechanism of action of histidyl-tRNAhis in regulating histidine gene expression. Seven contiguous histidine codons are present within this sequence: : formula: (see text)

Base Sequence↗