Learn More
We report ultrasensitive Ca(2+) indicators, yellow cameleon-Nano (YC-Nano), developed by engineering the Ca(2+)-sensing domain of a genetically encoded Ca(2+) indicator, YC2.60 or YC3.60. Their high Ca(2+) affinities (K(d) = 15-140 nM) and large signal change (1,450%) enabled detection of subtle Ca(2+) transients associated with intercellular signaling(More)
The structural modification of dendritic spines plays a critical role in synaptic plasticity. CaMKII is a pivotal molecule involved in this process through both kinase-dependent and independent structural functions, but the respective contributions of these two functions to the synaptic plasticity remain unclear. We demonstrate that the transient interplay(More)
All biological reactions depend on the diffusion and re-localization of biomolecules. Our understanding of biological processes requires accurate measurement of biomolecule mobility in living cells. Currently, approaches for investigating the mobility of biomolecules are generally restricted to measuring either fast or slow diffusion kinetics. We describe(More)
We report a pH-insensitive and photostable ultramarine fluorescent protein, Sirius, with an emission peak at 424 nm, the shortest emission wavelength among fluorescent proteins reported to date. The pH-insensitivity of Sirius allowed prolonged visualization of biological events in an acidic environment. Two fluorescence resonance energy transfer (FRET)(More)
Chromophore-assisted light inactivation (CALI) is a powerful technique for acute perturbation of biomolecules in a spatio-temporally defined manner in living specimen with reactive oxygen species (ROS). Whereas a chemical photosensitizer including fluorescein must be added to specimens exogenously and cannot be restricted to particular cells or sub-cellular(More)
We have used protein engineering to expand the palette of genetically encoded calcium ion (Ca(2+)) indicators to include orange and improved red fluorescent variants, and validated the latter for combined use with optogenetic activation by channelrhodopsin-2 (ChR2). These indicators feature intensiometric signal changes that are 1.7- to 9.7-fold improved(More)
For understanding the precise mechanisms of molecular recognition of proteins, three-dimensional structural analyses of the protein-protein complexes are essential. For this purpose, a new method to reveal complex structures was developed with the assistance of saturation transfer (SAT) and residual dipolar coupling (RDC) by heteronuclear NMR experiments,(More)
We have determined the binding site on agitoxin2 (AgTx2) to the KcsA K(+) channel by a transferred cross-saturation (TCS) experiment. The residues significantly affected in the TCS experiments formed a contiguous surface on AgTx2, and substitutions of the surface residues decreased the binding affinity to the KcsA K(+) channel. Based on properties of the(More)
The use of fluorescent proteins has revolutionized our understanding of biological processes. However, the requirement for external illumination precludes their universal application to the study of biological processes in all tissues. Although light can be created by chemiluminescence, light emission from existing chemiluminescent probes is too weak to use(More)
Chromophore-assisted light inactivation (CALI) is a potentially powerful tool for the acute disruption of a target protein inside living cells with high spatiotemporal resolution. This technology, however, has not been widely utilized, mainly because of the lack of an efficient chromophore as the photosensitizing agent for singlet oxygen ((1)O(2))(More)