Publications by authors named "Kazuki Shiraiwa"

2 Publications

  • Page 1 of 1

Ligand-directed two-step labeling to quantify neuronal glutamate receptor trafficking.

Nat Commun 2021 02 5;12(1):831. Epub 2021 Feb 5.

Department of Biomolecular Engineering, Graduate School of Engineering, Nagoya University, Nagoya, 464-8603, Japan.

The regulation of glutamate receptor localization is critical for development and synaptic plasticity in the central nervous system. Conventional biochemical and molecular biological approaches have been widely used to analyze glutamate receptor trafficking, especially for α-amino-3-hydroxy-5-methyl-4-isoxazole-propionate-type glutamate receptors (AMPARs). However, conflicting findings have been reported because of a lack of useful tools for analyzing endogenous AMPARs. Here, we develop a method for the rapid and selective labeling of AMPARs with chemical probes, by combining affinity-based protein labeling and bioorthogonal click chemistry under physiological temperature in culture medium. This method allows us to quantify AMPAR distribution and trafficking, which reveals some unique features of AMPARs, such as a long lifetime and a rapid recycling in neurons. This method is also successfully expanded to selectively label N-methyl-D-aspartate-type glutamate receptors. Thus, bioorthogonal two-step labeling may be a versatile tool for investigating the physiological and pathophysiological roles of glutamate receptors in neurons.
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http://dx.doi.org/10.1038/s41467-021-21082-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7864911PMC
February 2021

Chemical Tools for Endogenous Protein Labeling and Profiling.

Cell Chem Biol 2020 08 16;27(8):970-985. Epub 2020 Jul 16.

Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyoto 615-8510, Japan; JST-ERATO, Hamachi Innovative Molecular Technology for Neuroscience, Kyoto University, Kyoto 615-8530, Japan. Electronic address:

Protein analysis under biological conditions is now regarded as indispensable for understanding the structure and function of proteins, in addition to in vitro studies using purified target proteins. Because there are many molecules other than the protein-of-interest (POI) under live cell conditions, selective labeling of a POI is critical to distinguish the POI from other proteins for precise analysis. Protein labeling strategies utilizing genetically encoded tags have been used in POI modification in the complex environment of live cells. However, genetic manipulation may often induce overexpression of the POI and/or perturb the cellular context, resulting in unexpected artifacts in the protein analysis. Alternatively, recent progress in chemical biology has produced two major chemical approaches for analyzing endogenous proteins under native conditions. In this review, we summarize these techniques that utilize either protein-selective chemical labeling or proteome-directed chemical modification.
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http://dx.doi.org/10.1016/j.chembiol.2020.06.016DOI Listing
August 2020