A thermal denaturation study of chromatin and nuclease-produced chromatin fragments

1977 ◽  
Vol 55 (7) ◽  
pp. 736-746 ◽  
Author(s):  
Peter N. Lewis
Keyword(s):  
Thermal Denaturation ◽  
Micrococcal Nuclease ◽  
Melting Transition ◽  
Cleavage Sites ◽  
Main Determinant ◽  
Calf Thymus ◽  
Sulfhydryl Oxidation ◽  
Thermal Denaturation Study ◽  
Lower Temperature ◽  
Mechanical Shearing

Calf thymus chromatin and nuclease-produced chromatin fragments have been examined by thermal denaturation measurements. Native chromatin gave a series of distinct melting transitions at 64, 73, 79, and 85 °C in 0.25 mM EDTA pH 8. Treatments such as dialysis, mechanical shearing, or sulfhydryl oxidation of histone H3 carried out on native chromatin significantly altered the derivative melting profiles by blurring the distinct transitions and shifting the highest melting transition to a lower temperature. Derivative melting profiles for electrophoretically purified chromatin fragments, monomer through hexamer, all resembled that obtained from dialyzed chromatin. These results suggest that higher order structures exist in chromatin that are easily disrupted. Since the products of micrococcal nuclease (EC 3.1.4.7) digestion of the altered chromatins did not exhibit any major electrophoretic differences from those obtained from nuclei, then most likely the primary arrangements of histones along the DNA are the main determinant for cleavage sites.

Rearrangements and reconstitution of mononucleosomes as monitored by thermal denaturation measurements

1980 ◽  
Vol 58 (1) ◽  
pp. 73-81 ◽  
Author(s):  
Susan S. Chiu ◽  
Kay P. Lee ◽  
Peter N. Lewis
Keyword(s):  
Freeze Drying ◽  
Thermal Denaturation ◽  
Guanidine Hydrochloride ◽  
Melting Transition ◽  
Melting Profile ◽  
Base Pairs ◽  
Structural Domains ◽  
Calf Thymus ◽  
Dna Size ◽  
Histone Core

Derivative melting profiles of calf thymus mononucleosomes have been examined for changes resulting from variations in solvent pH and ionic strength, histone H1 content, and DNA size. Samples of mononucleosomes were found to rearrange during freeze-drying to form an altered monomer and a series of noncovalent multimers. The derivative melting profiles of these particles differ significantly from those for the untreated monomer and dimer. The noncovalent dimer exhibited a new melting transition at 66 °C involving approximately 18 base pairs of DNA normally associated with the highest melting transition. Mononucleosomes were reconstituted from 6 M guanidine hydrochloride to give particles with physical properties including melting profile which were virtually indistinguishable from those of the starting material. This result confirms the notion that no structural domains exist in the histone core that can be irreversibly denatured by noncovalent perturbations.

Solid-phase synthesis and thermal denaturation study of cyclic PNAs targeting the HIV-1 TAR RNA loop

2007 ◽  
Vol 17 (21) ◽  
pp. 6026-6030 ◽  
Author(s):  
Gregory Upert ◽  
Mohamed Mehiri ◽  
Audrey Di Giorgio ◽  
Roger Condom ◽  
Nadia Patino
Keyword(s):  
Thermal Denaturation ◽  
Solid Phase Synthesis ◽  
Solid Phase ◽  
Phase Synthesis ◽  
Thermal Denaturation Study ◽  
Tar Rna ◽  
Hiv 1

Plant 5S rDNA has multiple alternative nucleosome positions

Genome ◽  
2006 ◽  
Vol 49 (7) ◽  
pp. 840-850 ◽  
Author(s):  
Jaroslav Fulnecek ◽  
Roman Matyasek ◽  
Ales Kovarik
Keyword(s):  
Tandem Repeats ◽  
5S Rdna ◽  
Nicotiana Sylvestris ◽  
Micrococcal Nuclease ◽  
Dna Curvature ◽  
Cleavage Sites ◽  
Computer Prediction ◽  
Theoretical Computer ◽  
Nontranscribed Spacer ◽  
Multiple Alternative

In plants, 5S ribosomal DNA (5S rDNA) is typically found in hundreds of copies of tandemly arranged units. Nucleotide database searches revealed that the majority of 5S genes (>90%) have repeat lengths that are not simple multiples of a plant nucleosomal unit, ranging in plants from 175–185 bp. To get insight into the chromatin structure, we have determined positions of nucleosomes in the Nicotiana sylvestris and Nicotiana tomentosiformis 5S rDNA units with repeat lengths of about 430 and 645 bp, respectively. Mapping experiments carried out on isolated nucleo somal DNA revealed many (>50) micrococcal nuclease cleavage sites in each class of repeats. Permutation analysis and theoretical computer prediction showed multiple DNA bend sites, mostly located in the nontranscribed spacer region. The distance between bend sites, however, did not correspond to the average spacing of nucleosomes in 5S chromatin (~180 bp). These data indicate that 5S rDNA does not have fixed nucleosomal positioning sites and that units can be wrapped in a number of alternative nucleosome frames. Consequently, accessibility of transcription factors to cognate motifs might vary across the tandem array, potentially influencing gene expression.Key words: Nicotiana, 5S rDNA, heterochromatin, tandem repeats, nucleosomes, DNA curvature.

Influence of thermal denaturation on the electrophoretic mobility of calf thymus DNA

Biopolymers ◽  
1964 ◽  
Vol 2 (1) ◽  
pp. 1-8 ◽  
Author(s):  
L. Costaxtixo ◽  
A. M. Liquori ◽  
V. Vitagliaxo
Keyword(s):  
Electrophoretic Mobility ◽  
Thermal Denaturation ◽  
Calf Thymus Dna ◽  
Calf Thymus

scholarly journals FURTHER STUDIES OF INFECTIOUS DNA EXTRACTED FROM MYCOBACTERIOPHAGES

1966 ◽  
Vol 123 (2) ◽  
pp. 327-340 ◽  
Author(s):  
Margret I. Sellers ◽  
Tohru Tokunaga
Keyword(s):  
Mycobacterium Smegmatis ◽  
Base Composition ◽  
Thermal Denaturation ◽  
Buoyant Density ◽  
Plaque Formation ◽  
Double Stranded Dna ◽  
Calf Thymus ◽  
Phage Dna ◽  
Dna Thermal Denaturation ◽  
Adenine Thymine

Results of the previous investigation in which it was found that DNA extracted from D29 mycobacteriophage was infectious for Mycobacterium smegmatis 607, have been extended. DNA extracted from mycobacteriophage D4 and D32 produced plaques when plated on their respective hosts; D28 DNA, extracted in the same manner and tested under similar conditions, failed to show infectivity. Species barriers were not crossed by mycobacteriophage DNA; bacteria resistant to intact phage were not infected with the phage DNA. The efficiency of plating of the DNA is very much lower than that of intact phage; infection of a given host was not accomplished by DNA when titration for plaque formation by the intact phage was less than 109 PFU. The base composition of DNA extracted from the four mycobacteriophages and the three propagating hosts was very similar. The bases were paired, adenine with thymine and guanine with cytosine. A relatively higher per cent of guanine-cytosine than of adenine-thymine, was found. The buoyant density of each DNA in CsCl was linearly related to its guanine-cytosine content whereas with the exception of D28 DNA, thermal denaturation temperatures failed to show this relationship. However, the thermal transition profiles were characteristic of double stranded DNA. Additional evidence that D29 DNA forms complexes with basic proteins was obtained. Binding between calf thymus histone and between RNAase and D29 DNA readily occurs with a resultant loss in DNA infectivity. Trypsin and D29 DNA are only weakly reactive.

A study of the localization of high mobility group proteins in chromatin

1978 ◽  
Vol 56 (6) ◽  
pp. 480-491 ◽  
Author(s):  
Beatriz Levy W. ◽  
Gordon H. Dixon
Keyword(s):  
Mouse Brain ◽  
High Mobility ◽  
Cell Types ◽  
Dnase I ◽  
High Mobility Group ◽  
Micrococcal Nuclease ◽  
Hmg Proteins ◽  
Calf Thymus ◽  
Limited Digestion ◽  
Active Genes

High mobility group (HMG) proteins from fetal calf thymus and mouse brain chromatin were purified and compared electrophoretically. The four major HMG proteins characteristic of fetal calf thymus chromatin (HMG's 1, 2, 14, and 17) were also found to be present in mouse brain chromatin.Nuclei from these two eucaryotic tissues were digested with DNase I and micrococcal nuclease and the acid-soluble proteins solubilized by the two nucleases in both tissues were analyzed on starch gels.Limited digestion of fetal calf thymus nuclei with DNase I led to the solubilization of a substantial fraction of proteins HMG-1 and HMG-2 together with smaller amounts of H1. In addition, limited digestion with micrococcal nuclease released approximately 70% of HMG's 1 and 2 and variable amounts of H1 into the soluble fraction. The observation that HMG proteins 1 and 2 are selectively solubilized under conditions in which active genes have been shown to be preferentially digested in various other cell types suggests their selective association with chromatin regions which are transcriptionally competent.

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