Keiichi Inoue 교수

Light-driven outward H⁺ pumps are widely distributed in nature, converting sunlight energy into proton motive force. Here we report the characterization of an oppositely directed H⁺ pump with a similar architecture to outward pumps. A deep-ocean marine bacterium, Parvularcula oceani, contains three rhodopsins, one of which functions as a light-driven inward H⁺ pump when expressed in Escherichia coli and mouse neural cells. Detailed mechanistic analyses of the pur

Schizorhodopsins (SzRs), a rhodopsin family first identified in Asgard archaea, the archaeal group closest to eukaryotes, are present at a phylogenetically intermediate position between typical microbial rhodopsins and heliorhodopsins. However, the biological function and molecular properties of SzRs have not been reported. Here, SzRs from Asgardarchaeota and from a yet unknown microorganism are expressed in <i>Escherichia coli</i> and mammalian cells, and ion transport assays and patch clamp an

Muscarinic cholinergic receptor stimulation evokes catecholamine secretion from some cell types, but the mechanism has not been well characterized. Using pheochromocytoma (PC12) cells, we show that the muscarinic agonist methacholine stimulates 45Ca2+ influx and [3H]norepinephrine release in a dose-dependent manner. Experiments performed in Na+-free medium or with inhibitors of voltage-dependent Ca2+ channels suggest the involvement of a receptor-activated Ca2+ channel which differs significantl

Microbial rhodopsins are photoreceptive membrane proteins that transport various ions using light energy. While they are widely used in optogenetics to optically control neuronal activity, rhodopsins that function with longer-wavelength light are highly demanded because of their low phototoxicity and high tissue penetration. Here, we achieve a 40-nm red-shift in the absorption wavelength of a sodium-pump rhodopsin (KR2) by altering dipole moment of residues around the retinal chromophore (KR2 P2

In addition to the well-known light-driven outward proton pumps, novel ion-pumping rhodopsins functioning as outward Na(+) and inward Cl(-) pumps have been recently found in eubacteria. They convert light energy into transmembrane electrochemical potential difference, similar to the prototypical archaeal H(+) pump bacteriorhodopsin (BR) and Cl(-) pump halorhodopsin (HR). The H(+), Na(+), and Cl(-) pumps possess the conserved respective DTE, NDQ, and NTQ motifs in the helix C, which likely serve

There are two types of membrane-embedded ion transport machineries in nature. The ion pumps generate electrochemical potential by energy-coupled active ion transportation, while the ion channels produce action potential by stimulus-dependent passive ion transportation. About 80% of the amino acid residues of the light-driven proton pump archaerhodopsin-3 (AR3) and the light-gated cation channel channelrhodopsin (ChR) differ although they share the close similarity in architecture. Therefore, the

Thousands of light-driven proton-pumping rhodopsins have been found in marine microbes, and a light-driven sodium-ion pumping rhodopsin was recently discovered, which utilizes sunlight for the energy source of the cell. Similarly, a light-driven chloride-ion pump has been found from marine bacteria, and three eubacterial light-driven pumps possess the DTE (proton pump), NDQ (sodium-ion pump), and NTQ (chloride-ion pump) motifs corresponding to the D85, T89, and D96 positions in bacteriorhodopsin

Sodium-pumping rhodopsins (NaRs) are light-driven outward Na(+) pumps. NaRs have a conserved Asn, Asp, and Gln motif (NDQ) in the third transmembrane helix (helix C). The NDQ motif is thus expected to play a crucial role in the operation of the Na(+) pump. Herein, we studied the photocycles of the NDQ-motif mutants of Krokinobacter rhodopsin 2 (KR2), the first discovered NaR, by flash photolysis, to obtain insight into the mechanism of Na(+) transport. For example, the KR2 N112A mutant did not a

The diffusion coefficients (D) of poly-L-glutamic acid (PLG) at various pHs are investigated by the laser-induced transient-grating method with a new photoreactive probe molecule. The pH dependence of D is compared with that of the helical content of PLG measured by circular dichroism. It is found that the pH dependences of both quantities are very similar. Since the frictions of the translational diffusion of charged and protonated carboxyl groups are found to be similar each other, it is concl