3P285 CheY-P binding to the bacterial flagellar motor affects not only the direction but also the speed of rotation(26. Measurements,Poster)

Abstract

Quantitative modelling has become an essential procedure toward understanding the cell at the systems level. Insights into the inner workings of biological networks can be obtained from mathematical analysis of biochemical models that might not be possible from direct experimentation alone. E-Cell System is one of the cell simulation platforms that allow us model cellular biochemical reaction networks in single-molecule resolution. For further quantitative understanding of intracellular molecular dynamics and kinetics, we have developed fluorescence microscopy/ spectroscopy Monte Carlo simulation to have direct comparison of the simulated data to single molecule experimental data. 3P285細菌べん毛モーターへの CheY-P の結合は回転方向だけでな く速度にも影響する。 Many bacteria swim in liquid environments by rotating helical flagella by an ion-driven rotary motor. The motor rotates in the counter-clockwise (CCW) direction constantly and in the clockwise (CW) direction occasionally for smooth swimming and tumbling, respectively. The CW rotation occurs by the binding of phosphorylated CheY (CheY-P) to the motor as the signal molecule. The behavior and specification of the motor have been well studied in the CCW rotation but little is known about the CW rotation. We carried out bead-rotation assay on a FliG mutant motor that rotates only in the CW direction without CheY-P and found that the motor function is symmetrical in the CCW and CW rotation. We also found that CheY-P binding to the motor disturbs the motor function. Isotope effect in the conventional neutron protein crystallography (NPC) can be eliminated by proton polarization technique (ppt). Furthermore, the ppt can make hydrogen visibility about 8 times larger than that in the conventional NPC, etc. On the other hand, several technical difficulties should be overcome in order to perform the ppt. In this presentation, several fundamental studies to realize ppt will be presented; the high pressure flash cooling was found a promising technique to make bulk water amorphous, and an ESR measurement will be presented in order to detect the spin distribution in a protein single crystal which is important to polarize protons in a protein. 3P287 Ｘ線自由電子レーザーを利用した球状生体超分子複合体のコ ヒーレントＸ線イメージングへの取り組みApproaches to cohherent X-ray diffraction imaging of single virus particle using X-ray free-electron laser The advent of X-ray free-electron leasers (X-FEL) creates new capabilities for coherent x-ray diffraction imaging (CXDI) of a single macromolecule. The first image reconstruction of a biological molecule using CXDI techniques was performed by Seibert et al. (2011). Low-resolution images of mimi-virus were calculated from the single diffraction pattern using X-FEL from the Linac Coherent Light Source (LCLS) at Stanford. In Japan, an X-FEL facility, SACLA (SPring-8 Angstrom Compact Free Electron Laser) has been developed, which produces high brilliance and short X-ray pulses at Angstrom wavelength. The use of the facility began with the start of user operation in March, 2012. We obtained some diffraction images...