[3P-2] Poster 3 (3P-48ー3P-87)

[3P-72] Single-molecule study of conformational dynamics of Intrinsically disordered E. coli Cytidine Repressor DNA Binding domain

^○Shrutarshi Mitra¹, Hiroyuki Oikawa^1,2, Athi N. Naganathan³, Satoshi Takahashi^1,2 (1.Department of Chemistry, Tohoku University, 2.Institute for Multidisciplinary Research for Advanced Materials, Tohoku University, 3.Department of Biotechnology, Bhupat & Jyoti Mehta School of Biosciences, Indian Institute of Technology, Madras)

DNA-binding domain of E. coli cytidine repressor (CytR) is intrinsically disordered but folds upon binding to its target. Although previous studies revealed a large array of conformations in the absence of DNA, a single-molecule approach could give quantitative insight into the role of individual microstates in the specific DNA binding. Herein we chose the alternating laser excitation scheme of smFRET as an experimental tool, which is capable of measuring accurate FRET based on 2-D Stoichiometry-Efficiency histogram. Our initial investigations at different ionic strengths and in denaturant like GdmCl showed that CytR populates multiple conformations at low ionic strength but converts to a single compact form at higher ionic strength and other way around expands gradually in GdmCl. Upon the association with the target and non-specific DNA, CytR converts into the compact form similar to the salt-induced structure, suggesting electrostatic potential of DNA drives the binding of CytR in a non-specific manner. These observations indicate that CytR, despite significant disordering, possesses a fair extent of folded-like state and interconverts on a very slow timescale (~ms) but in the presence of DNA coalesces to a folded form rapidly.