11:00-12:00 Controlling the Revolving and Rotating Motion Direction of Asymmetric Hexameric Nanomotor by Arginine Finger and Channel Chirality
Peixuan Guo,
College of Pharmacy
Many ATPases are nanoscale motion machines that are classified into three categories based on the motion mechanisms in transporting substrates: linear, rotating, and the recently discovered revolving motor (see here). Many ATPases are composed of multi-subunit ring-shaped structures that hydrolyze ATP to generate forces. An intriguing question is how these biomotors control the motion direction and regulate the sequential action of their multiple subunits. Many, but not all, ATPases are hexamers containing a conserved arginine finger in each subunit to control the direction of motion and coordinating the interactions of adjacent subunits. We use crystal structure, Cryo-EM, single molecule TIRF, optical tweezers, biotechnology, ATP analogue blockage and other biophysical approaches to illuminate the mechanisms of the motion direction control, inter-subunit interactions and sequential movements of individual subunits. The conclusion was supported by the of asymmetrical hexamer structure with one dimer and four monomers in the high-resolution structures of many ATPases. The trans-acting arginine residue is situated at the interface of two subunits and extends into the ATP binding pocket of the downstream subunit. Channel chirality and channel size are distinct between revolving or rotating biomotors. The conformational changes and entropy alternation triggered by ATP binding and hydrolysis will be presented as a new mechanism that deviates from the traditional concept of ATP-mediated mechanochemical energy coupling. The elucidation of motor structure, motion mechanism and direction control of polygon-shaped ATPases could provide a prototype for the fabrication of new types of motors in nanotechnology or at macro level.
Further reading:
12:20-1:20 Chromatin higher order folding, the physics of DNA organization
John van Noort,
Leiden University
Our genome is densely packed in a dynamic, hierarchical fashion, balancing compaction with controlled regulation of DNA accessibility. DNA organization starts with strings of nucleosomes, 147 bp DNA segments that are wrapped twice around 8 histone proteins, and fold into dense chromatin fibers. However, the structure of chromatin fibers is poorly defined and heavily debated. We used single-molecule techniques to probe and manipulate the dynamics of nucleosomes in individual chromatin fibers. These novel methods were initially applied to synthetic, highly homogeneous nucleosomal arrays. We found a strong dependence of the structure of the fibers on the length of the linker DNA, which was corroborated by rigid basepair Monte Carlo simulations. Unfortunately, synthetic chromatin lacks the complexity that provides functionality to our epi-genome. We recently developed a method to purify specific chromatin fragments from yeast without crosslinking the fiber. Magnetic Tweezers based force spectroscopy on intact, native fibers uniquely probes chromatin structure, composition and variations in it at the single-molecule level. Though we observed reduced unfolding forces, the native fibers showed similar stiffness and unfolding pathways as compared to synthetic chromatin. Our systematic single-molecule analysis of a wide range of chromatin compositions supports a general picture of nucleosomes stacking in 1- and 2-start topologies, whose stability is determined by the length of the linker DNA. These experimental results constrain the wide range of chromatin models and bring us closer to ab initio prediction of higher order chromatin folding.