Atomic force microscopic study of aggregation of RecA-DNA nucleoprotein filaments into left-handed supercoiled bundles.
RecA and its complexes with double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA) are responsible for homologous recombination and DNA repair. In this study, we have observed, by atomic force microscopy (AFM), two-filament left-handed superhelices of RecA-dsDNA filaments that further interwind into four- or six-filament bundles, in addition to previously reported left-handed bundles of three or six filaments. Also revealed are four-filament bundles formed by further interwinding of two intrafilament superhelices of individual filaments. Pitches of superhelices of RecA-DNA filaments are similar to each other regardless the number of component filaments, and those formed on Phix174 RFII dsDNA and pNEB206A dsDNA are measured as 339.3 +/- 6.2 nm (690 counts of pitch/2) and 321.6 +/- 11.7 nm (101 counts of pitch/2), respectively, consistent with earlier measurements made by electron microscopy with a much smaller sample size. The study of these structures provides insight into the self-interactions of RecA and RecA-like proteins, which are present in all living cells, and into the general phenomenon of bundling, which is relevant to both biological and nonbiological filaments.
Shi
Wei-Xian
WX
Department of Chemical Engineering, University of Michigan, 3074 H.H. Dow, 2300 Hayward Street, Ann Arbor, MI 48109-2136, USA.
Larson
Ronald G
RG
D001665:Binding Sites; D003460:Crystallization / Q000379:methods*; D004247:DNA / Q000737:chemistry* / Q000648:ultrastructure*; D007536:Isomerism; D008422:Materials Testing; D018625:Microscopy, Atomic Force; D046912:Multiprotein Complexes / Q000737:chemistry; D049329:Nanostructures / Q000737:chemistry* / Q000648:ultrastructure*; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation; D011938:Rec A Recombinases / Q000737:chemistry* / Q000648:ultrastructure*
D046912:Multiprotein Complexes; D004247:DNA; D011938:Rec A Recombinases