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T Niidome

Publications and source records attributed to T Niidome.

67 records · Page 4Linked to original sources
PubMed1993

Molecular cloning of the smaller subunit(P52) of rat liver mitochondrial processing protease.

A cDNA encoding the smaller subunit (P52) of mitochondrial processing protease was isolated from a rat liver cDNA library, using cDNA fragment for yeast MAS1 as the probe. The deduced amino acid sequence is highly homologous to those of PEP from Neurospora crassa and MAS1 from Saccharomyces cerevisiae. After in vitro transcription and translation, the precursor peptide was imported into isolated rat liver mitochondria and processed to its mature form.

Amino Acid Sequence↗
PubMed1993

Protease D from the sarcocarp of honeydew melon fruit.

A serine protease was purified from commercially available honeydew melon fruit (Cucumis melo L. var. inodorus Naud) by five steps including CM-Sepharose CL-6B column chromatography. At the first CM-Sepharose CL-6B chromatography four active fractions appeared. No difference in enzymatic properties and M(r) in each fraction was found. Finally, honeydew melon protease D was isolated from the major active fraction D. A M(r) of 50,000 was determined for protease D on SDS-PAGE and gel filtration. Protease D was a glycoprotein. The protease exhibited a remarkable stability, even in 8 M urea or 2 M guanidine hydrochloride for 24 hr. The optimum pH of the enzyme was 11 at 35 degrees using casein as substrate. The enzyme was strongly inhibited by diisopropyl fluorophosphate and was not inhibited by metal-chelating reagents or cysteine protease inhibitors. The enzymatic properties of honeydew melon protease D were similar to those of cucumisin [EC 3.4.21.25], except for its stability at high temperature, wide pH range and against detergents.

Amino Acid Sequence↗
PubMed1993

Primary structure and functional expression of the omega-conotoxin-sensitive N-type calcium channel from rabbit brain.

The complete amino acid sequence of a rabbit brain calcium channel (BIII) has been deduced by cloning and sequencing the cDNA. The open reading frame encodes 2339 amino acids, which corresponds to an M(r) of 261,167. A phylogenetic tree representing evolutionary relationships indicates that BIII is grouped together with the other rabbit brain calcium channels, BI and BII, into a subfamily that is distinct from the dihydropyridine-sensitive L-type subfamily. Transient expression in cultured skeletal muscle myotubes derived from muscular dysgenic mice demonstrates that the BIII channel mediates an omega-conotoxin-sensitive calcium current with kinetics and voltage dependence like those previously reported for whole-cell N-type current. Cell-attached patch recordings, with isotonic barium as the charge carrier, revealed distinct single channels with an average slope conductance of 14.3 pS.

Amino Acid Sequence↗
PubMed1992

Function of a truncated dihydropyridine receptor as both voltage sensor and calcium channel.

The skeletal muscle dihydropyridine (DHP) receptor serves dual functions, as a voltage sensor for excitation-contraction coupling and as an L-type calcium channel. Biochemical analysis indicates the presence of two forms of the DHP receptor polypeptide in skeletal muscle, a full-length translation product present as a minor species and a much more abundant form that has a truncated carboxy-terminus. On the basis of these and other observations, it has been proposed that, in skeletal muscle, only the full-length DHP receptor can function as a calcium channel and that the truncated form can only function as a voltage sensor for excitation-contraction coupling. To resolve this issue, we have now constructed a complementary DNA (pC6 delta 1) encoding a protein corresponding to the truncated DHP receptor in skeletal muscle. Expression of pC6 delta 1 in dysgenic myotubes fully restores both excitation-contraction coupling and calcium current, consistent with the idea that a single class of DHP receptors performs both functions.

Amino Acid Sequence↗
PubMed1992

Molecular cloning and characterization of a novel calcium channel from rabbit brain.

The complete amino acid sequence of a novel calcium channel (designated BII) from rabbit brain has been deduced by cloning and sequencing the cDNA. The BII calcium channel is structurally more closely related to the BI calcium channel than to the cardiac and skeletal muscle L-type calcium channels. Blot hybridization analysis of RNA from different tissues and from different regions of the brain shows that the BII calcium channel is distributed predominantly in the brain, being abundant in the cerebral cortex, hippocampus and corpus striatum.

Amino Acid Sequence↗
PubMed1990

Regions of the skeletal muscle dihydropyridine receptor critical for excitation-contraction coupling.

It is thought that in skeletal muscle excitation-contraction (EC) coupling, the release of Ca2+ from the sarcoplasmic reticulum is controlled by the dihydropyridine (DHP) receptor in the transverse tubular membrane, where it serves as the voltage sensor. We have shown previously that injection of an expression plasmid carrying the skeletal muscle DHP receptor complementary DNA restores EC coupling and L-type calcium current that are missing in skeletal muscle myotubes from mutant mice with muscular dysgenesis. This restored coupling resembles normal skeletal muscle EC coupling, which does not require entry of extracellular Ca2+. By contrast, injection into dysgenic myotubes of an expression plasmid carrying the cardiac DHP receptor cDNA produces L-type calcium current and cardiac-type EC coupling, which does require entry of extracellular Ca2+. To identify the regions responsible for this important functional difference between the two structurally similar DHP receptors, we have expressed various chimaeric DHP receptor cDNAs in dysgenic myotubes. The results obtained indicate that the putative cytoplasmic region between repeats II and III of the skeletal muscle DHP receptor is an important determinant of skeletal-type EC coupling.

Animals↗
PubMed1990

Purification and characterization of an acidic amino acid-specific endopeptidase of Bacillus subtilis obtained from a commercial preparation (Protease Type XVI, Sigma).

An acidic amino acid-specific endopeptidase was purified from Protease Type XVI (Sigma), a commercial product from culture filtrate of Bacillus subtilis, by a series of column chromatographies on CM-Toyopearl (Fractogel) and Mono-S, guided by activity assay using Boc-Ala-Ala-Pro-Glu-pNA as a substrate. The final preparation was homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and reversed-phase HPLC. The molecular weight of the protease was estimated to be 18,000 by gel filtration on TSK gel G3000SWXL column using 6 M guanidine hydrochloride as an eluent, and 17,000 by SDS-PAGE in the presence of 2-mercaptoethanol. The isoelectric point of the protease was 7.7. Studies on the substrate specificity with peptide p-nitroanilides and natural peptides revealed that the protease hydrolyzes the peptide bonds on the carboxyl-terminal side of acidic amino acids, especially of glutamic acid. The protease was completely inactivated by DFP, indicating the serine protease nature of the protease. The activity of the protease was also inhibited by EDTA and GEDTA, and reactivated by Ca2+. The protease contained 1.3 +/- 0.2 mol/mol protein of Ca2+. These results suggest that Ca2+ plays a vital role in the protease activity.

Amino Acid Sequence↗
PubMed1989

Primary structure and functional expression from complementary DNA of the rod photoreceptor cyclic GMP-gated channel.

The complete amino-acid sequence of the cyclic GMP-gated channel from bovine retinal rod photoreceptors, deduced by cloning and sequencing its complementary DNA, shows that the protein contains several putative transmembrane segments, followed by a region that is similar to the cyclic GMP-binding domains of cyclic GMP-dependent protein kinase. Expression of the complementary DNA produces cyclic GMP-gated channel activity in Xenopus oocytes.

Amino Acid Sequence↗
PubMed1989

Primary structure and functional expression of the cardiac dihydropyridine-sensitive calcium channel.

In cardiac muscle, where Ca2+ influx across the sarcolemma is essential for contraction, the dihydropyridine (DHP)-sensitive L-type calcium channel represents the major entry pathway of extracellular Ca2+. We have previously elucidated the primary structure of the rabbit skeletal muscle DHP receptor by cloning and sequencing the complementary DNA. An expression plasmid carrying this cDNA, microinjected into cultured skeletal muscle cells from mice with muscular dysgenesis, has been shown to restore both excitation-contraction coupling and slow calcium current missing from these cells, so that a dual role for the DHP receptor in skeletal muscle transverse tubules is suggested. We report here the complete amino-acid sequence of the rabbit cardiac DHP receptor, deduced from the cDNA sequence. We also show that messenger RNA derived from the cardiac DHP receptor cDNA is sufficient to direct the formation of a functional DHP-sensitive calcium channel in Xenopus oocytes. Furthermore, higher calcium-channel activity is observed when mRNA specific for the polypeptide of relative molecular mass approximately 140,000 (alpha 2-subunit) associated with skeletal muscle DHP receptor is co-injected.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMedDate not supplied

Chain length of cationic alpha-helical peptide sufficient for gene delivery into cells.

To define the minimal peptide length needed for gene delivery into mammalian cells, we synthesized several peptides with shortened chain lengths from the amino-termini of the original amphiphilic peptides (4(6), Ac-LARL-LARL-LARL-LRAL-LRAL-LRAL-NH( 2,) and Hel 11-7, KLLK-LLLK-LWKK-LLKL-LK), which have been known to have gene transfer abilities into cells. Each synthetic peptide was studied for its ability to bind and aggregate with plasmid DNA and the structural change of the peptide caused by binding with the DNA to establish a relative in vitro gene transfection efficiency in COS-7 cells. As a result, the deletion of eight amino acid residues of 4(6) had little influence on their ability, whereas that of 12 amino acid residues remarkably reduced the abilities to make aggregates and transfer the DNA into the cell. In the case of the Hel 11-7 series peptides, deletion of amino acid residues caused a considerable reduction in abilities to bind and form aggregates with DNA and to transfer the DNA into cell in due order. In summary, 16 and 17 amino acid residues were sufficient to form aggregates with the DNA and transfer the DNA into the cells in the deletion series of 4(6) and Hel 11-7, respectively. Furthermore, it was indicated that reduction of membrane perturbation activity of the peptide-DNA complex due to deletion of the peptide chain length caused suppression of the transfection efficiency even if the complex was incorporated into the cells. Transfer of the complex to cytosol mediated by membrane perturbation activity of the peptide is an important step for efficient protein expression from its cDNA. The results of this study will make it easy to design and synthesize a functional gene carrier molecule such as a carbohydrate-modified peptide used in targeted gene delivery.

Amino Acid Sequence↗