AFM for analysis of structure and dynamics of DNA and protein–DNA complexes

Abstract

This paper describes protocols for studies of structure and dynamics of DNA and protein–DNA complexes with atomic force microscopy (AFM) utilizing the surface chemistry approach. The necessary specifics for the preparation of functionalized surfaces and AFM probes with the use of silanes and silatranes, including the protocols for synthesis of silatranes are provided. The methodology of studies of local and global conformations DNA with the major focus on the time-lapse imaging of DNA in aqueous solutions is illustrated by the study of dynamics of Holliday junctions including branch migration. The analysis of nucleosome dynamics is selected as an example to illustrate the application of the time-lapse AFM to studies of dynamics of protein–DNA complexes. The force spectroscopy is the modality of AFM with a great importance to various fields of biomedical studies. The AFM force spectroscopy approach for studies of specific protein–DNA complexes is illustrated by the data on analysis of dynamics of synaptic SfiI–DNA complexes. When necessary, additional specifics are added to the corresponding example.