An analysis of transcription factor TFIIIA-mediated DNA supercoiling.

We have analyzed transcription factor-mediated DNA supercoiling catalyzed by the Xenopus oocyte extract (S-150). Under conditions that inhibit endogenous supercoiling activity (2 mM EDTA), the 5S RNA specific transcription factor, TFIIIA, promotes a negative change in DNA linking number. The SV40 binding protein, T antigen, appears not to promote DNA supercoiling under these conditions.

Restoration of the anterior neck surface in the burned patient by free groin flap.

In treating extensive burn contractures of the anterior neck, we obtained good results using free flaps. Ninety-nine patients with neck contractures were treated with free flaps. The ages of the patients range from 2 to 64 years.

Synthesis and properties of oligodeoxyribonucleotides containing abasic site.

Dodecanucleotide, d(CGCXCGCGTGCG), containing abasic site at desired position in the sequence was synthesized by solid-phase triester method. The introduced moiety (X) WAS N-(2'-deoxy-beta-D-erythropentofuranosyl) formamide.

Cloning, sequencing and genetic mapping of a Bacillus subtilis cell wall hydrolase gene.

We have cloned DNA fragments from Bacillus subtilis 168S into Escherichia coli, which produced a lytic zone on an agar medium containing B. subtilis cell wall. Sequencing of the fragments showed the presence of an open reading frame (ORF) which encodes a polypeptide of 272 amino acids with a molecular mass of 29919 Da.

Transcription factor TFIIIA stimulates DNA supercoiling promoted by a fractionated cell-free extract from Xenopus laevis.

An activity that can introduce negative supercoils into relaxed covalently closed DNA molecules has been isolated from a Xenopus laevis cell-free extract (S-150) and purified over 200-fold. The exogenous addition of ATP, other ribonucleotides and deoxyribonucleotides, as well as nonhydrolyzable analogs, stimulate DNA supercoiling which may occur by a pathway involving multiple protein components. DNA supercoiling occurs in topological single steps and is inhibited by camptothecin and berenil, but not novobiocin or VM-26, suggesting a catalytic role for topoisomerase I in the reaction.

[A study of linearity and reciprocity during shock applied with a hammer to human dry skull].

The authors used a human dry skull on which the cranial bone mandible had been joined with an artificial articulator disk to form a single unit. Impact acceleration corresponding to weak and strong tapping was considered a dynamic load in examining the vibration transfer characteristics of the facial cranial bone when impact was applied from the mentum section in a situation designed to be closer to reality. Flexion injection type (resonance frequency f0 = 100 to 150 Hz, produced by GC Corp.

Ultrastructural localization of fibronectin and laminin in human granulation tissue in relation to capillary development.

The distribution of fibronectin (FN) and laminin (LM) at developing capillaries during various developmental stages, from capillary sprouts to relatively developed capillaries, was studied by light- and electron-microscopy immunocytochemistry. By light-microscope, FN immunoreactivity was diffusely distributed throughout the stroma of the granulation tissues, while for LM it was preferentially distributed at the perivascular region with the various developmental stages of the immature capillaries. Ultrastructural study revealed that capillary sprouts were closely surrounded by plentiful deposits of immunoreaction with the FN, but only faintly for LM.

Nucleotide sequences of the Bacillus subtilis flaD locus and a B. licheniformis homologue affecting the autolysin level and flagellation.

A DNA fragment containing the flaD locus of Bacillus subtilis, which had been cloned into plasmid pAC3, was subcloned into an M13 phage and sequenced. The sequence contained five open reading frames (ORFs), of which ORF2 was the flaD gene. Unexpectedly, the sequence of the flaD locus was identical to that of sin [sporulation inhibition gene; Gaur, N.

cis-acting enhancement of RNA polymerase III gene expression in vitro.

The Xenopus laevis S-150 cell-free extract catalyzes in vitro transcription of several RNA polymerase III genes. Among these are the Xenopus 5S RNA gene (somatic type) and the Xenopus methionine tRNA gene. In this report we present an analysis of the transcriptional activity of these two genes either in trans-competition experiments or when the genes are co-localized in the same circular plasmid.

Characterization and comparison of mitochondrial DNAs and rRNAs from Penicillium urticae and P. chrysogenum.

Mitochondrial DNA (mt DNA) from a patulin producer, Penicillium urticae (synonym P. griseofulvum), was 27.8 kb +/- 0.

Reaction parameters of TFIIIA-induced supercoiling catalyzed by a Xenopus laevis cell-free extract.

In addition to its fundamental role of nucleating the formation of stable transcription complexes, the Xenopus laevis 5S RNA specific transcription factor, TFIIIA, promotes a variety of DNA-associated metabolic reactions. We report that TFIIIA can induce a DNA supercoiling catalyzed by the Xenopus laevis S-150 cell-free extract on plasmids containing a single copy of the Xenopus 5S RNA gene (somatic-type). Stimulated supercoiling occurs in the presence of high concentrations of ATP (4 mM) and at a factor to DNA ratio of 1 through a mechanism most likely involving type I topoisomerase.

Interactions between quadruple-stranded oligodeoxyribonucleotides and drugs.

Circular dichroism (CD) and fluorescence spectra have been measured for complexes formed between four-stranded G4-DNA and ethidium bromide (EB). The EB-G4-DNA complexes showed similar induced CD spectra, compared with the induced CD spectrum of the EB-calf thymus DNA complex.

DNA superhelicity enhances the assembly of transcriptionally active chromatin in vitro.

Using an in vitro chromatin assembly system, we analyzed the influence of DNA superhelicity on the development of transcriptionally active minichromosomes. Plasmid DNA molecules containing either a Xenopus borealis 5S RNA gene or an X. laevis methionine tRNA gene were utilized as templates for the assembly of chromatin.

Studies on the ATP requirements of in vitro chromatin assembly.

To gain a more complete understanding of the process of in vitro chromatin assembly, an examination of the energy requirements of nucleosome formation must be undertaken. The experiments outlined in this manuscript address this issue by making use of the Xenopus laevis S-150 cell-free extract. The S-150 catalyzes chromatin assembly on circular DNA templates dependent either on the exogenous addition of ATP or regeneration of endogenous ATP.

Changes in DNA topology can modulate in vitro transcription of certain RNA polymerase III genes.

The role of DNA supercoiling in eukaryotic gene expression is not fully understood. The objective of this study was to examine the regulation of in vitro chromatin assembly by topological alterations in the DNA template using a cell-free extract from Xenopus laevis oocytes (S-150). The results suggest that input DNA topology may be a determining factor in controlling the transcriptional activity of the Xenopus tRNA and one particular 5S gene.

Self-assembly of synthetic deoxyoligonucleotides containing d-G clusters.

Deoxyoligonucleotides containing d-G cluster were synthesized to elucidate exact structures and properties of the parallel-four stranded complexes of oligo(dG). Among these oligomers, d-TTGGGGTT and d-TTGGGGGGTT formed stable complexes which were able to interact with ethidium bromide known as an intercalator.

Studies on DNA topoisomerase activity during in vitro chromatin assembly.

The in vitro assembly of chromatin, promoted by the Xenopus cell-free extract (S-150), can be inhibited by oxolinic acid and to a lesser extent by nalidixic acid. Both of these antibiotics have been shown to block the activity of the specialized type II Topoisomerase, bacterial DNA Gyrase. Oxolinic acid induces a DNA cleavage by Micrococcal Nuclease at specific sequences in the multiple cloning vector pGEM-4.

Molecular cloning of a gene affecting the autolysin level and flagellation in Bacillus subtilis.

A 2.8 kb PstI fragment of Bacillus subtilis 168W DNA has been cloned into Escherichia coli HB101 and B. subtilis AG5 using pAC3 as a shuttle plasmid.

6-Methyl-1,2,4-benzenetriol, a new intermediate in penicillic acid biosynthesis in Penicillium cyclopium.

Penicillic acid-negative mutants were obtained from a color mutant derived from Penicillium cyclopium NRRL 1888 through N-methyl-N'-nitro-N-nitrosoguanidine treatment. One mutant (SK2N6) accumulated 6-methyl-1,2,4-benzenetriol, which was not previously known to be a metabolite of P. cyclopium, in addition to orsellinic acid and orcinol.

Genetic mapping by means of protoplast fusion in Bacillus subtilis.

A new mapping method involving protoplast fusion in Bacillus subtilis is described. Protoplasts from an isogenic standard marker strain containing purA and from a strain containing both purB and the marker, "x", to be mapped were fused with polyethylene glycol, and purA+ purB+ fusants were selected. After isolation of single colonies and determination of unselected markers, marker x was mapped between two standard markers.

Characterization of chromosome and plasmid transformation in Bacillus subtilis using gently lysed protoplasts.

Competent cells of Bacillus subtilis were transformed with DNA from gently lysed protoplasts. Significant linkages among markers separated by distances of approximately 2.3% of the total chromosome were found, which have not been detected for conventional transformation.

New macrolides from Penicillium urticae mutant S11R59.

Two new macrolides, patulolide B and patulolide C, were isolated from a culture filtrate of Penicillium urticae S11R59 mutant. The structures of these macrolides were determined and their biological activities were investigated. These structures and biological activities were also compared with those of patulolide A which was produced by the same organism.

Selection methods in bacilli for recombinants and transformants of intra- and interspecific fused protoplasts.

The intraspecific fusion frequencies obtained with the direct selection method on a semi-synthetic regeneration medium between strains of B. subtilis and B. licheniformis were distributed from 9.