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At least 91 records · Page 5Linked to original sources
PubMed2001

Kinetic proofreading can explain the supression of supercoiling of circular DNA molecules by type-II topoisomerases.

The enzymes that pass DNA through DNA so as to remove entanglements, adenosine-triphosphate-hydrolyzing type-II topoisomerases, are able to suppress the probability of self-entanglements (knots) and mutual entanglements (links) between approximately 10 kb plasmids, well below the levels expected, given the assumption that the topoisomerases pass DNA segments at random by thermal motion. This implies that a 10-nm type-II topoisomerase can somehow sense the topology of a large DNA. We previously introduced a "kinetic proofreading" model which supposes the enzyme to require two successive collisions in order to allow exchange of DNA segments, and we showed how it could quantitatively explain the reduction in knotting and linking complexity. Here we show how the same model quantitatively explains the reduced variance of the double-helix linking number (supercoiling) distribution observed experimentally.

Binding Sites↗
PubMed1974

Locating interrupted hydrogen bonding in the secondary structure of PM2 circular DNA by comparative denaturation mapping.

Previous studies with HCHO have revealed a reaction with superhelical DNA that strongly suggests that this DNA consists of small regions of interrupted secondary structure. To map these sites in PM2 DNA, the following set of experiments was performed using electron microscopy. (i) A denaturation map of nicked form II was obtained using Inman's alkaline-HCHO conditions. (ii) The superhelical form I was reacted with HCHO at 30 C until equilibrium was achieved at the interrupted sites (3.6% reactivity). The excess HCHO was removed rapidly and X-ray treatment was employed to nick these prereacted molecules. These form II molecules containing HCHO (form II HCHO) were also subjected to denaturation mapping. It would be expected that the HCHO-unpaired regions would serve as induction sites for the propagation of melting. Hence, depending on the location of the induction sites; we would anticipate either the creation of new regions of melting or a normal denaturation map shifted to lower pH values. Comparison of the development of progressive denaturation of form II and form II HCHO reveals that the latter is the case. The denaturation maps of form II are highly organized patterns of adenine-thymine (AT)-rich regions, with a total of five regions at extreme pH conditions. There are six highly organized regions for form II HCHO, i.e., smaller adjacent loops, at low denaturation conditions where no denaturation is seen for form II. These coalesce into the pattern for form II containing four of five A-T-rich regions observed for form II. Hence we conclude that the regions of altered hydrogen bonding in superhelical PM2 DNA are four to six in number and they map in the A-T-rich regions of the DNA.

Bacteriophages↗
PubMed1980

Molecular cloning of the Harvey sarcoma virus circular DNA intermediates. II. Further structural analyses.

Three species of unintegrated supercoiled Harvey sarcoma virus DNA (6.6, 6.0, and 5.4 kilobase pairs) have been molecularly cloned from Harvey sarcoma virus-infected cells. On the basis of restriction enzyme analyses, the 6.6- and 6.0-kilobase pair viral DNAs contain two and one copies, respectively, of a 650-base pair DNA segment which contains sequences present at the 3' and 5' termini of the viral genome. R-loop structures formed between Moloney leukemia virus RNA and the cloned Harvey sarcoma virus DNA indicated that about 500 base pairs of the 650-base pair repeating segment was complementary to the 3' end of the viral RNA. During amplification in the Escherichia coli host, some recombinants containing the 6.6- or the 6.0-kilobase pair Harvey sarcoma virus DNA insert acquired or lost the complete 650-base pair DNA segment. These changes occurred in both recA+ and recA- E. coli.

Animals↗
PubMedDate not supplied

[Analysis of the sequence of repeats in divergent regions of maxi-circular DNA from kinetoplasts of Crithidia oncopelti].

The nucleotide sequence of direct clustered repeats from the divergent region of the maxicircle kinetoplast (mitochondrial) DNA from the protozoan Crithidia oncopelti was analysed. 10 kbp long divergent region contains 3 clusters composed of 18-23 tandem repeats of 82-83 bp (Sau-repeats) and a single cluster of five 417 bp repeats (EcoRI-repeats). The clusters are interspersed between the regions of nonrepetitive DNA. The details of the structural organization of repeats and their clusters were considered on the nucleotide sequence level. The possible ways of origin of the clusters are discussed.

Animals↗
PubMed2003

An apparatus for electrophoretic capture and recovery of circular DNA in thin layers.

An apparatus was designed for the electrophoretic capture and recovery of circular DNA in thin layers (membranes). Rapid separations were done by the use of a low-conductivity buffer and high electric field strengths. Two methods that specifically retain circular DNA in the membranes were demonstrated using the supercoiled and open circular forms of two plasmids with sizes of 4.4 and 13 kbp. Electrophoretic trapping (by an impalement mechanism) in agarose-gel-filled membranes used electric field strength to immobilize circular DNA in the membranes. The other method of capture utilized the greatly reduced electrophoretic mobility of circular DNA in membranes composed of agarose and the linear polymer hydroxyethylcellulose. The reduction in electrophoretic mobility was not dependent upon the electric field strength, distinguishing it from electrophoretic trapping. Trapping of circular DNA in the membranes followed by size analysis using agarose-gel electrophoresis could be used as a two-dimensional separation tool for the analysis of complex mixtures. Captured DNA was recovered by two methods: (i). centrifugation of membranes made with low-melting-point agarose resulted in a gel slurry that could be heated to release the DNA; (ii). electroelution of the membranes. Electroelution was done by using an electrode that was isolated from the DNA by a 'barrier' membrane. A non-trapping electric field was used to reverse the DNA out of the membrane into a small volume of buffer above the membrane.

Adsorption↗