The Interactions Between SSB Protein And SsDNA Studies By Using AFM

Nucleic acids and proteins are two important biomacromolecules that compose the life, and the interactions between them are the base of many biological phenomena, such as growth, propagation, inheritance, metabolism, and so on. The nano-mechanical detection of nucleic acid-protein interactions at single-molecule level will deepen our understanding and eventually gain controls on these biological processes. This is also the key to explore the mysteries of life. Atomic force microscopy (AFM) imaging can directly show the mode of intra (or inter)-molecular interactions, and AFM-based single molecule force spectroscopy (SMFS) can be used to measure the strength of intra (or inter)-molecular interactions quantitatively. In our research, firstly, we synthesized a kind of long single-stranded DNA (ssDNA) with multiply repeated copies, and then let it hybridize with short complementary DNA fragments forming the nicked double-stranded DNA (dsDNA). The force-extension curves of pure ssDNA as well as the nicked dsDNA were obtained by using SMFS. Secondly, the long chain ssDNA were allowed to interact with native SSB proteins from Bacillus subtilis the binding mode of the ssDNA-SSB complexes were characterized by using imaging function of AFM. Then we used the long ssDNA as a probe to detect the effect of SSB binding on the nanomechanical properties of ssDNA. Quantitative binding energy between ssDNA and SSB protein was obtained by comparing the difference of mechanic properties between pure ssDNA and SSB-ssDNA complexes.