Atomic force microscopy of nucleoprotein complexes.

Recent data on the AFM studies of nucleoprotein complexes of different types are reviewed in this paper. The first section describes the progress in the sample preparation methods for AFM studies of nucleic acids and nucleoprotein complexes. The second part of this paper reviews AFM data on studies of complexes of DNA with regulatory proteins. These studies include two different types of DNA distortion induced by proteins binding: local bending of DNA at sites of protein binding and formation of large loops due to protein-protein interactions between molecules bound to distant sites along the DNA molecules (DNA looping). The prospects for use of AFM for physical mapping of genomes are discussed in this section as well. The third part of the paper reviews data on studies of complexes of DNA with non-sequence specific binding proteins. Special emphasis is given to studies of chromatin which have resulted in progress in the understanding of structure of native chromatin fiber. In this section, novel data on AFM studies of RecA-DNA filaments and complexes of dsRNA with the dsRNA-specific protein p25 are also presented. Discussion of the substrate preparation procedures in relation to the AFM studies of nucleoprotein complexes is given in the final section

Characterizing nanoscale morphologic and mechanical properties of α-synuclein amyloid fibrils with atomic force microscopy

We have used atomic force microscopy (AFM) to image wild-type and disease-related mutant α-synuclein (aS) amyloid fibrils deposited on various hard and soft surfaces, ranging from freshly cleaved mica to two different supported lipid bilayers: phosphatidylcholine (POPC) and a mixture of POPC and phosphatidylglycerol (POPG). Quantitative morphologic analyses show that fibrils deposited on freshly cleaved mica, gold and glass substrates appeared to have similar heights and lengths, suggesting that the surface–fibril interaction in these cases does not influence the fibril morphology. When the same fibril sample is deposited on HOPG or quartz, the aS fibrils appear shorter in length and completely distorted, respectively, indicating that the interaction with these surfaces severely affects the fibril morphology. Sequentially recorded AFM images of fibrils on POPC bilayers clearly revealed that the amyloid fibrils were mobile on the bilayer, indicating that the fibrils are in thermal equilibrium on the surface. Persistence lengths of the fibrils on mica and POPC have been determined using two different methods, and reveal no significant differences, indicating that, although the fibrils appear immobile on mica, they do thermally equilibrate in 2D before they finally attach to the substrate. The persistence length of aS fibrils is found to be between 3.3 and 7.1 μm, which is within the expected range for amyloid fibrils. We observe no fibril movement on the mixed POPC/POPG bilayer, suggesting a stronger interaction with the lipids. Furthermore, the reduced heights of the fibrils on top of the mixed bilayer suggest that the fibrils are partly embedded within the bilayer

Surface characterization using atomic force microscopy (AFM) in liquid environments

Liquid imaging provides intrinsic advantages for AFM experiments, particularly for conducting in situ studies of chemical or biochemical reactions. Using liquid media has benefits for improving resolution, since the amount of force applied between the tip and sample can be reduced. Surface changes caused by immersion in different liquids can be investigated, such as for studying electrochemical reactions with different parameters of solvent polarity, pH or ion concentration. Aqueous buffers enable studies of biochemical reactions that simulate physiological conditions, with time-lapse capture of image frames at different intervals. Studies of surface changes throughout the course of self-assembly reactions have been monitored with AFM in liquid media. By injecting new molecules into the sample cell, AFM-based nanofabrication can be accomplished by nanografting protocols. Liquid environments expand the capabilities for scanning probe studies to provide insight for dynamic processes at the molecular-level. © Springer-Verlag Berlin Heidelberg 2013