SUNY at Albany
June 19-23, 2001
Ultrastructural studies of circular superhelical DNA complexes with trivaline
Intramolecular compact structures formed by circular superhelical DNA molecules of different molecular weights due to interaction with synthetic oligopeptide trivaline were studied by atomic force and electron microscopy. Several DNA preparations were used: lower molecular weight plasmids pBl, pLg-19 with sizes 2920 bp, 3820 bp and higher molecular weight plasmids pTbo1, pRX10 and cosmid 27877, with sizes 6120 bp, 10500 bp and 44890 bp respectively. Plasmid pTbo1 and pRX10 preparations along with monomers contained significant amount of dimers and trimers. Main structures in all preparations observed were compact particles, which coincide in their appearance and compaction coefficient (3,5-3,7) with triple rings described earlier. The size and structure characteristics of triple rings and other compact particles on atomic force images in general coincide with those obtained by EM. AFM images allow to get additional information about the ultrastructural organization and arrangement of DNA fibers within the compact structures. Along with triple rings in pTbo1 and pRX10 Ð TVP complexes significant amount of compact structures were observed having the shape of two or three compact rings attached to each other by a region of compact fibre. Basing on the data of contour length measurements and the shape of the particles it was concluded that these structures were formed due to compaction of dimeric and trimeric circular DNA molecules. Structures consisting of several attached to each other triple rings were not found for any of monomeric plasmid or cosmid preparations Ð TVP complexes where only single triple rings were observed. The conclusion is made that initiation of compact fibre formation within the circular molecules depends on the primary structure and for dimeric or trimeric circular molecules two or three compaction initiation points are present, located in each monomer unit within one circular DNA molecule. Basing on the ultrastructural data obtained the models of DNA fibers organization in all kinds of compact structures observed are proposed. The nucleotide sequence dependent compaction mechanism providing independent compaction of portions of one circular molecule can be of interest for understanding of DNA compaction processes in vivo.
Yuri Yu. Vengerov (1), Larissa P. Martinkina (1), Dmitry V. Klinov (2), Alexander A. Kolesnikov (3), Vyacheslav Yu. Yurchenko (3), Sergey A. Streltsov (1) Tatyana V. Neretina (2), Victor V. Demin (2)
Engelhardt Institute of Molecular Biology(1), Russian Academy of Sciences, Vavilov str., 32, Moscow, 119991 Russia(2), Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Mikluho-Maklaya 16/10, Moscow, 117871 Russia(3), Lomonossov Moscow State University, Department of Biology, Moscow, 119899 Russia,