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Biomedical subjects

H Probst

Publications and source records attributed to H Probst.

At least 73 records · Page 4Linked to original sources
PubMed1978

Differences in the binding of methylated albumin to non-replicating, replicating and denatured DNA from Ehrlich ascites cells.

The binding of methylated albumin to DNA, the basis of the chromatography on columns of kieselguhr coated with methylated albumin (MAK chromatography), was investigated. Scatchard plots revealed only one mode of interaction with fully double-stranded DNA. The complexes should be completely dissociated by raising the NaCl concentration of the solution to 0.8 M, indicating a binding by electrostatic attraction between the oppositely charged protein and DNA molecules. In complexes with denatured and partially single-stranded replicating DNA an additional kind of binding was found which made these complexes more stable against salt dissociation. These secondary interactions were stronger at 23 degrees C than at 0 degrees C and could be weakened by the addition of 6 M urea. It was therefore concluded that apolar forces were involved in these interactions.

Animals↗
PubMed1976

Newly synthesized mammalian cell DNA. Separation of Okazaki pieces by thermal chromatography on hydroxyapatite.

The optical melting profile of a fraction of nascent DNA which can be isolated from DNA of logarithmically growing Ehrlich ascites cells by nitrocellullose column chromatography was recorded. In relation to the bulk DNA a slightly lowered Tm and an increased width of the transition was observed. Thermal hydroxyapatite chromatography revealed that melting of the nascent DNA begins at relatively low temperatures in helix regions containing small daughter strand pieces. The pieces were characterized as Okazaki pieces by determining their size and their labeling kinetics during [3H] thymidine pulses. The combination of nitrocellulose chromatography and thermal hydroxyapatite chromatography seems to be suitable for isolating Okazaki pieces.

Animals↗
PubMed1975

[Repair replication in synchronous Ehrlich ascites cells after reversal of a methotrexate block (author's transl)].

If synchronous Ehrlich ascites cells are arrested at the beginning of the S phase by a methotrexate block, and the block is reversed by addition of thymidine to the culture, the cells incorporate a considerable amount of this thymidine into parental DNA within the first minute. These newly synthesized segments are irregularly distributed among the total parental DNA. This kind of distribution is expected if these segments are the result of excision repair. On the basis of the kinetics of the process this repair probably is DNA synthesis in regions already excised during the methotrexate block.

Animals↗
PubMed1975

[The secondary structure and nitrocellulose affinity of freshly replicated DNA from Ehrlich ascites cells (author's transl)].

Hydroxyapatite chromatography and isopycnic Cs2SO4 centrifugation normally yield no indications of single-stranded DNA when that fraction of replicating DNA from Ehrlich ascites cells which can be separated by nitrocellulose chromatography is analyzed. Single-stranded DNA is detected by both methods if the DNA is fragmented by ultrasound before the nitrocellulose chromatography. The digestion of this DNA fraction by single-strand-specific nucliase leads to a loss of its binding to nitrocellulose and of the indications of single-stranded DNA. The loss for the affinity to nitrocellulose is also observed when the corresponding fraction separated from unfragmented DNA is digested by endonuclease. It is suggested that replicating DNA is bound to nitrocellulose by means of single-stranded gaps on the replication fork. These gaps are apparently too small to be detected within large, otherwise entirely double-stranded molecules by hydroxyapatite chromatography and Cs2SO4 centrifugation. In the case of nitrocellulose-binding ultrasound fragments, this relation seems to be more favorable because of the separation of most of the residual double-stranded part. It is demonstrated that sonication of helical DNA also generates a small amount of fragments with some single-stranded character. The effects observed with replicating DNA could be distinguished from these artifacts.

Animals↗