Biomedical subjects

Alexey Bogdanov

Publications and source records attributed to Alexey Bogdanov.

2 recordsLinked to original sources

PubMed2003

DNA recombination: holliday junctions dynamics and branch migration.

Holliday junctions are critical intermediates for homologous, site-specific recombination, DNA repair, and replication. A wealth of structural information is available for immobile four-way junctions, but the controversy on the mechanism of branch migration of Holliday junctions remains unsolved. Two models for the mechanism of branch migration were suggested. According to the early model of Alberts-Meselson-Sigal (Sigal, N., and Alberts, B. (1972) J. Mol. Biol. 71, 789-793 and Meselson, M. (1972) J. Mol. Biol. 71, 795-798), exchanging DNA strands around the junction remain parallel during branch migration. Kinetic studies of branch migration (Panyutin, I. G., and Hsieh, P. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 2021-2025) suggest an alternative model in which the junction adopts an extended conformation. We tested these models using a Holliday junction undergoing branch migration and time-lapse atomic force microscopy, an imaging technique capable of imaging DNA dynamics. The single molecule atomic force microscopy experiments performed in the presence and in the absence of divalent cations show that mobile Holliday junctions adopt an unfolded conformation during branch migration that is retained despite a broad range of motion in the arms of the junction. This conformation of the junction remains unchanged until strand separation. The data obtained support the model for branch migration having the extended conformation of the Holliday junction.

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PubMed2002

Crosslinking of 4.5S RNA to the Escherichia coli ribosome in the presence or absence of the protein Ffh.

Radioactively labeled 4.5S RNA containing statistically distributed 4-thiouridine residues in place of normal uridine was prepared by T7 transcription. The ability of this modified 4.5S RNA to form a complex with the protein Ffh was demonstrated by a gel shift assay. The modified 4.5S RNA, with or without Ffh, was added to Escherichia coli ribosomes under various conditions, and crosslinking from the thiouridine residues was induced by irradiation at 350 nm. The crosslinked ribosomal components were analyzed by our standard procedures. Two clearly defined types of crosslinking were observed. The first was a crosslink to 23S rRNA, which was entirely dependent both on the presence of Ffh and a nascent protein chain in the 50S subunit. This crosslink was localized to nt approximately 2828-2837 of the 23S rRNA, from position 84 of the 4.5S molecule. The second type of crosslinking, to the 30S ribosomal subunit, was independent of the presence of Ffh, and was found both with vacant 70S ribosomes or isolated 30S subunits. Here the crosslink was localized to the 3'-terminal region of the 16S rRNA, from positions 29-50 of the 4.5S RNA. Cross-linking to ribosomal protein S1 was also observed. The known crystal structure of the protein Ffh/4.5S RNA fragment complex was extrapolated by computer modeling so as to include the whole 4.5S molecule, and this was docked onto the ribosome using the crosslinking data. The results are discussed in terms of multiple functions and binding sites of the 4.5S RNA.

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