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19 results on '"Uno, Tomohide"'

16. Further Evaluation of the Localization and Functionality of Hemagglutinin Epitope- and Fluorescent Protein-Tagged AtMinD1 in Arabidopsis thaliana.

Author

Fujiwara, Makoto T., Dongliang Li, Kazama, Yusuke, Abe, Tomoko, Uno, Tomohide, Yamagata, Hiroshi, Kanamaru, Kengo, and Itoh, Ryuuichi D.

Subjects
ARABIDOPSIS thaliana, HEMAGGLUTININ, EPITOPES, PHENOTYPES, CHLOROPLASTS
Abstract

The article presents a study that examines the localization and functionality of Arabidopsis thaliana MinD (AtMinD1) fused with a dual hemagglutinin epitope (dHA) or a yellow fluorescent protein (YFP). It used the wildtype Arabidopsis thaliana to provide a basis of MinD function and localization in chloroplast division. Results say that AtMinD1-dHA successfully complements the arc11/atminD1 mutant phenotype and is located at the envelope membrane and the mid-chloroplast constriction site.

Published
2009
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17. Rice Bifunctional α-AmyIase/Subtilisin Inhibitor: Cloning anti Characterization of the Recombinant Inhibitor Expressed in Escherichia coli.

Author

Yamasaki, Teruyuki, Deguchi, Masaki, Fujimoto, Toshiko, Masumura, Takehiro, Uno, Tomohide, Kanamaru, Kengo, and Yamagata, Hiroshi

Subjects
MOLECULAR cloning, CLONING, ESCHERICHIA coli, GERMINATION, PLANT physiology, AMYLASES, AMYLASE inhibitors, DIGESTIVE enzymes
Abstract

The article presents a study that examines cloning and characterization of the recombinant inhibitor expressed in Escherichia coli. The possible role in the regulation of seed germination in the prevention of precocious germination had been discussed. Nucleotide sequence of the cDNA and its gene that encode a bifunctional rice amylase/subtillisin inhibitor (RASI) were analyzed. Based on the results, it has been suggested that RASI might function in the defense of the seed against microorganisms.

Published
2006
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18. Molecular Cloning of cDNA for BRab from the Brain of Bombyx moriand Biochemical Properties of BRab Expressed in Escherichia coli

Author

UNO, Tomohide, UENO, Mayumi, NAKAJIMA, Ayumi, SHIRAI, Yasuhito, and AIZONO, Yasuo

Abstract

From a brain cDNA library of Bombyx mori, we cloned cDNA for BRab, which encoded a 202-amino-acid polypeptide sharing 60-80% similarity with rab1 family members. To characterize its biochemical properties, cDNA for BRab was inserted into an expression vector (pGEX2T) and expressed in Escherichia colias a glutathione S-transferase (GST) fusion protein. The recombinant protein was purified to homogeneity with glutathione S-Sepharose. The purified GST-BRab bound [35S]-GTPγS and [3H]-GDP with association constants of 1.5×106M-1and 0.58×106M-1, respectively. The binding of [35S]-GTPγS was inhibited with GTP and GDP, but with no other nucleotides. The GTP-hydrolysis activity was evaluated to be 5 m mole/min/mole of BRab. In the presence of 6 mMMgCl2, bound [35S]-GTPγS and [3H]-GDP were exchanged with GTPγS most efficiently. These results suggest that BRab, having a higher affinity for GTP than GDP, converts from the GTP-bound state into the GDP-bound state by intrinsic GTP hydrolysis activity and returns to the GTP-bound state with the exchange of GDP with GTP.

Published
1998
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19. Rice bifunctional alpha-amylase/subtilisin inhibitor: cloning and characterization of the recombinant inhibitor expressed in Escherichia coli.

Author

Yamasaki T, Deguchi M, Fujimoto T, Masumura T, Uno T, Kanamaru K, and Yamagata H

Subjects
Amino Acid Sequence, Base Sequence, Cloning, Molecular, Escherichia coli genetics, Molecular Sequence Data, Oryza chemistry, Oryza metabolism, Plant Proteins genetics, Plant Proteins pharmacology, RNA, Messenger analysis, RNA, Messenger metabolism, Recombinant Proteins genetics, Recombinant Proteins pharmacology, Subtilisin antagonists & inhibitors, alpha-Amylases antagonists & inhibitors, Escherichia coli metabolism, Oryza genetics, Plant Proteins biosynthesis, Recombinant Proteins biosynthesis
Abstract

The complete nucleotide sequences of the cDNA and its gene that encode a bifunctional alpha-amylase/subtilisin inhibitor of rice (Oryza sativa L.) (RASI) were analyzed. RASI cDNA (939 bp) encoded a 200-residue polypeptide with a molecular mass of 21,417 Da, including a signal peptide of 22 amino acids. Sequence comparison and phylogenetic analysis showed that RASI is closely related to alpha-amylase/subtilisin inhibitors from barley and wheat. RASI was found to be expressed only in seeds, suggesting that it has a seed-specific function. A coding region of RASI cDNA without the signal peptide was introduced into Escherichia coli and was expressed as a His-tagged protein. Recombinant RASI was purified to homogeneity in a single step by Ni-chelating affinity column chromatography and characterized to elucidate the target enzyme. The recombinant inhibitor had strong inhibitory activity toward subtilisin, with an equimolar relationship, comparable with that of native RASI, and weak inhibitory activity toward some microbial alpha-amylases, but not toward animal or insect alpha-amylases. These results suggest that RASI might function in the defense of the seed against microorganisms.

Published
2006
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