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B Henrich

Publications and source records attributed to B Henrich.

49 records · Page 3Linked to original sources
PubMed1990

Peptidase D gene (pepD) of Escherichia coli K-12: nucleotide sequence, transcript mapping, and comparison with other peptidase genes.

The nucleotide sequence of a 2.3-kilobase-pair DNA fragment of Escherichia coli that contains the transcription signals and the coding region of the pepD gene specifying aminopeptidase D was determined. The location and extent of the open reading frame were verified by partial amino acid sequencing of the purified pepD product. By use of a promoter-screening vector, initiation signals for pepD transcription were located in the 5'-flanking region of the open reading frame. Analysis of pepD transcripts by S1 mapping, primer extension, and Northern (RNA) hybridization revealed two species of monocistronic mRNA with different 5' ends and a common 3' end. Calculation of the degree of codon usage bias in the coding region suggested that the efficiency of pepD translation is relatively low. As deduced from the predicted amino acid sequence, peptidase D is a slightly hydrophilic protein of 485 amino acid residues that contains no extended domains of marked hydrophobicity. Structural and functional features of the pepD gene are discussed and compared with other already sequenced peptidase genes of E. coli.

Amino Acid Sequence↗
PubMed1989

Accurate mapping of the Escherichia coli pepD gene by sequence analysis of its 5' flanking region.

A cloned DNA fragment, carrying the gene for peptidase D (pepD) of Escherichia coli, was partially sequenced. By purification of peptidase D and sequence determination of an amino-terminal oligopeptide the reading frame of the pepD gene, starting with a GTG initiator codon, was unambiguously identified. An overlap of the established nucleotide sequence with the previously sequenced 5' flanking region of the gpt gene allowed the exact distance between pepD and gpt to be calculated. The two genes are pointing towards each other and are separated by 260 bp. A search for open reading frames (ORFs) and the analysis of possible codon usage in the intercistronic region indicate the absence of an additional gene (lpcA) between pepD and gpt.

Amino Acid Sequence↗
PubMed1986

Use of the lysis gene of bacteriophage phi X174 for the construction of a positive selection vector.

DNA fragments generated by a variety of restriction enzymes can easily be cloned in the small (3.2-kb) positive-selection vector pUH84, which contains the modified lysis gene of bacteriophage phi X174 under transcriptional control of the lac promoter. Plasmid pUH84 does not yield transformants after introduction into Escherichia coli unless the lysis gene is inactivated by insertion of foreign DNA into one of the unique PstI, SalI, AccI, HincII, BamHI, or EcoRI sites. This highly efficient positive selection of recombinants requires neither the use of a distinct host strain nor a special induction of the lysis function. Transcription of fragments cloned into pUH84 may be effectively regulated by the lac promoter provided the host cells are cotransformed with the newly constructed plasmid pUH7 which carries the IQ allele of the lac repressor gene.

Bacteriophage phi X 174↗
PubMed1986

Cloning and expression of the pepD gene of Escherichia coli.

Peptidase D of Escherichia coli, cleaving the unusual dipeptide carnosine, was found to be encoded by the ColE1 hybrid plasmid pLC44-11. From this plasmid the pepD gene was subcloned into small vectors. As shown by successive reduction of the flanking sequences of genomic DNA, the order of genes in the region at 6 min of the E. coli K12 map is phoE, pepD, in the clockwise orientation. Insertional inactivation of the pepD gene and expression of recombinant plasmids in maxicells allowed the identification of the pepD product as a 52 kDa protein. Comparison with the 100 kDa protein molecular mass determined by gel filtration suggests that active peptidase D is probably a dimer.

Cloning, Molecular↗
PubMed1985

Lysis of Escherichia coli by cloned phi X174 gene E depends on its expression.

The lysis gene E of bacteriophage phi X174 was cloned under transcriptional control of the lefthanded lambda promoter, giving rise to plasmid pSB12. Plasmid pSB22, identical to pSB12 except for an amber mutation in gene E, was constructed in the same way. Induction of the cloned wild-type gene by heat inactivation of the thermosensitive lambda cI857 repressor resulted in lysis of the host bacteria. With plasmid pSB22 only amber suppressor strains of Escherichia coli lysed after heat inactivation of lambda cI857. Lysis of E. coli was shown to depend on the rate of gene E translation and on the growth phase of the bacteria. Stationary cells could not be lysed by the gene E product (gpE), even if present in sufficient amounts to lyse growing cells. By isotopic labelling gpE could be detected among the proteins synthesized in normal E. coli as well as in minicells. Determination of gene E expression suggested that gpE synthesis is translationally regulated.

Bacteriophage phi X 174↗
PubMed1985

Effect of iodide treatment on iodine concentration and volume of endemic non-toxic goitre in childhood.

In a total of 195 children and adolescents of both sex (mean age 12.9, range 5-17 years) with endemic non-toxic goitre the thyroidal iodine concentration (IC) was determined using X-ray fluorescent scanning on admission and during iodine (100 micrograms daily) and L-thyroxine (3 micrograms/kg body weight daily) treatment respectively. Additionally the thyroid volume was measured sonographically in a longitudinal study including 46 patients before and after 4-8 months of iodine supplementation (100 micrograms daily). The IC was 305 +/- 144 micrograms/g. It compared well with that of adult goitre patients (288 +/- 109 micrograms/g) and was significantly inferior to the value of normal controls (389 +/- 170 micrograms/g). Under L-thyroxine therapy the IC further decreased (243 +/- 144 micrograms/g), whereas patients receiving iodide showed an increase of the IC (570 +/- 197 micrograms/g). The mean TSH level fell from 2.3 +/- 0.9 microU/ml to 1.4 +/- 0.6 microU/ml. The average T4/TBG (thyroxine binding globulin) ratio showed a slight increase which, however, was not significant. The mean goitre volume decreased by 40%. It was evidenced that iodide is useful not only in the prophylaxis of non-toxic goitre but also as a more physiologic treatment than thyroid hormones, at least for young subjects with simple diffuse goitres.

Adolescent↗
PubMed1984

Use of a cloned bacteriophage gene to disrupt bacteria.

A plasmid, pUH51, was constructed, which contains the lysis gene E of bacteriophage phi X174, subjected to the regulatory region of the lac operon, as well as the lac repressor gene. This plasmid can readily replicate in any strain of E. coli and mediates lysis of the bacteria after induction of the cloned phi X174 gene E. Taking advantage of these properties, plasmid pUH51 was used as a tool for gentle disruption of E. coli. At cell concentrations below 5 X 10(10)/ml, the efficiency of this method, as measured by release of beta-galactosidase from the cells, exceeded the efficiency of conventional methods for cell breakage.

Alleles↗
PubMed1982

Lysis of Escherichia coli by induction of cloned phi X174 genes.

The largest of the fragments produced by AluI digestion of phi X174 RFI DNA comprises genes E and J as well as parts of genes D and F. This DNA fragment (1007 bp) was cloned into the lac z' gene of plasmid pUR222. In the recombinant plasmid pUH12, transcription of the phi X174 genes is controlled by the lac p-o region. Induction of the cloned genes by addition of the lac inducer, IPTG, resulted in lysis of the bacteria. Cloning of the corresponding AluI-fragment from phi X174am3 DNA, carrying an amber mutation in gene E, showed that the expression of this gene alone is sufficient to trigger cell lysis. The time interval between the addition of IPTG and the onset of lysis depended on the concentration of the inducer, however, the rate of lysis was similar at all IPTG concentrations used.

Bacteriophage phi X 174↗
PubMed1980

Demonstration of the early--late switch in vitro with bacteriophage T7 DNA as template.

A protein-synthesizing system in vitro, programmed with bacteriophage T7 DNA as template, changed the specificity of gene expression in the course of incubation as a result of newly synthesized T7 early proteins. The system mimics largely the situation in vivo on both the transcriptional and the translational levels, i.e. early gene expression is turned off shortly after late synthesis has been started. These results suggest that the switch from early to late expression does not necessarily require changes in the cellular environment nor is it dependent on the presence of membranes. The main part in this process is played by the phage-dependent RNA polymerase (gene 1 product), whose activity appears 8-10 min after start of incubation. When its activity is reduced by inhibitors or creation of non-optimal conditions, the system is not able to manage the early--late switch.

DNA, Viral↗
PubMedDate not supplied

Cloning and expression of P60, a conserved surface-localized protein of Mycoplasma hominis, in Escherichia coli.

The clp60 gene encoding P60, a conserved lipoprotein of Mycoplasma hominis, was cloned and sequenced from both the type strain PG21 and the isolate FBG. Both open reading frames were identical in length, comprising 1746 nucleotides. The deduced amino acid sequences differed in 16 out of 582 amino acids. As expected, none of these divergences mapped within the epitope that was recognized by mAb CG4 in all of the 198 isolates of M. hominis analyzed so far. This conserved epitope was narrowed down to amino acids 454 through 464 within the C terminus of P60. For the expression of the recombinant homolog P60, P60rec, in E. coli the TGA codons of clp60 were substituted for TGG codons prior to cloning of clp60 into the expression plasmid pQE41. The expression of P60rec as a fusion protein with dihydrofolate reductase carrying an N-terminal His-tag enabled the purification of large amounts of P60rec in a soluble form.

Amino Acid Sequence↗