In Vivo Assembly and Arrangement of the DNA Translocase SpoIIIE During Chromosome Segregation and Membrane Fission in B. Subtilis

Abstract

lumen represents a continuous nanofluidic network for the transport of proteins and small molecules throughout the cells. Here, we describe a system in which synthetic interconnected, millimeter-scale nanofluidic networks were formed from the cooperative interaction between phospholipid vesicles and motor protein-based transport. In this system, the energy-driven transport of microtubule filaments by kinesin motors provides a "pulling force" that acts on multilamellar liposomes, connected via biotin-streptavidin bonds, and results in the formation of highly bifurcated networks of lipid nanotubes. Moreover, the processing of microtubules by the motors enables self-healing within this system in which nanotubes are continuously elongating and collapsing while maintaining the overall network morphology. The size/shape of the networks can be further modulated by regulating microtubule surface density, as well as total amount and physical properties of the lipid. Once formed, we characterized the diffusivity of the lipids and the ability of these networks to support materials transport. Attachment of quantum dots to lipids within the networks suggests that the lipids remain highly fluid, and that particle transport closely follows a onedimensional process when the particle density is low, and a single-file, onedimensional process when particle density is high. Current work is focused on characterizing the connectivity of multi-vesicle networks and interstitial materials transport as a model system for studying biomolecular transport and communication. Sandia National Laboratories is a multi-program laboratory managed and operated by Sandia Corporation, a wholly owned subsidiary of Lockheed Martin DNA is an interesting building block for the construction of active nanoscale devices. Movement in these structures is typically achieved through conformational changes that can be driven by DNA fuel strands or by internal changes in DNA hybridization. In this study we investigate the function and fidelity of a DNA actuator with 11 states using single molecule FRET microscopy. The DNA actuator is a complex molecular DNA device capable of finely controlled actuation through the movement of its two DNA piston arms, see the included figure. The actuator is designed to have 10 actuation steps of less than 1 nm each and to be locked in a desired state by the addition of specific locking strands. Our results show that the DNA actuator can be effectively locked in place in several conformations with the help of well-designed DNA locking strands. However, depending on the sequence of the locking strands, the device can also show spontaneous oscillations between welldefined states. Our results demonstrate that single molecule studies provide a stringent test for the performance of molecular machines made out of DNA. In all three domains of life, Sm oligomers interact with RNA, either as chaperones or as scaffolding to facilitate RNA$$$RNA and RNA$$$protein interactions. In bacteria and eukarya, Sm homologs are key for RNA metabolism, f...