title: Deinococcus radiodurans RecA nucleoprotein filaments characterized at the single-molecule level with optical tweezers.
abstract: Deinococcus radiodurans can survive extreme doses of ionizing radiation due to the very efficient DNA repair mechanisms that are able to cope even with hundreds of double-strand breaks. RecA, the critical protein of homologous recombination in bacteria, is one of the key components of the DNA-repair system. Repair of double-strand breaks requires RecA binding to DNA and assembly of the RecA nucleoprotein helical filaments. The Escherichia coli RecA protein (EcRecA) and its interactions with DNA have been extensively studied using various approaches including single-molecule techniques, while the D. radiodurans RecA (DrRecA) remains much less characterized. However, DrRecA shows some remarkable differences from E. coli homolog. Here we combine microfluidics and single-molecule DNA manipulation with optical tweezers to follow the binding of DrRecA to long double-stranded DNA molecules and probe the mechanical properties of DrRecA nucleoprotein filaments at physiological pH. Our data provide a direct comparison of DrRecA and EcRecA binding to double-stranded DNA under identical conditions. We report a significantly faster filaments assembly as well as lower values of persistence length and contour length for DrRecA nucleoprotein filaments compared to EcRecA. Our results support the existing model of DrRecA forming more frequent and less continuous filaments relative to those of EcRecA.
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia. Electronic address: george.pobegalov@nanobio.spbstu.ru.
authors.forename: Georgii
authors.initials: G
authors.lastname: Pobegalov
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Galina
authors.initials: G
authors.lastname: Cherevatenko
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Aleksandr
authors.initials: A
authors.lastname: Alekseev
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Anton
authors.initials: A
authors.lastname: Sabantsev
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Oksana
authors.initials: O
authors.lastname: Kovaleva
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Alexey
authors.initials: A
authors.lastname: Vedyaykin
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Natalia
authors.initials: N
authors.lastname: Morozova
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia; Petersburg Nuclear Physics Institute, NRC Kurchatov Institute, Gatchina 188300, Russia.
authors.forename: Dmitrii
authors.initials: D
authors.lastname: Baitin
authors.affiliation: Peter the Great St.Petersburg Polytechnic University, Saint-Petersburg 195251, Russia.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Khodorkovskii
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001696:Biomechanical Phenomena; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D034301:Deinococcus / Q000378:metabolism* / Q000528:radiation effects; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D046210:Microfluidic Analytical Techniques; D009698:Nucleoproteins / Q000737:chemistry / Q000378:metabolism; D052898:Optical Tweezers; D055503:Protein Multimerization; D011938:Rec A Recombinases / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D009698:Nucleoproteins; D011938:Rec A Recombinases
keywords: Deinococcus radiodurans; Homologous recombination; Optical tweezers; RecA