Visualization of dynamins

The tools presented in this chapter have been used to characterize the structural and biochemical properties of dynamins. The versatility in microscopic techniques allows for visualization of dynamins with varied shapes, including ring, spiral, and helical oligomers. Negative stain allows for structures to be examined quickly; however, larger structures may flatten, as was observed with Dnm1. Cryo-EM helps eliminate flattening, as shown with Dnm1, allows the specimen to be viewed in a more native state and freezing the sample rapidly following substrate addition (i.e., GTP) allows conformational changes to be observed in seconds. In addition, AFM, fluorescence microscopy, and light scattering assays can be used to observe structural rearrangements that occur upon GTP hydrolysis. Rotary shadowing reveals the surface structure of the sample and can be used to determine the hand of the helical array. The accuracy of STEM analysis provides a means to calculate atomic mass over a defined area or length of helix. Together, these methods provide a comprehensive approach for visualizing dynamins. © 2008 Elsevier Inc. All rights reserved.