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Biomedical subjects

M Doi

Publications and source records attributed to M Doi.

At least 163 records · Page 9Linked to original sources
PubMed1995

Calbindin-D 28k immunoreactivity in the cerebellum of spinocerebellar degeneration.

We studied immunoreactivity for calbindin-D 28k (CaBP), an intracellular calcium-binding protein, in the cerebellum of control subjects and of patients with spinocerebellar degeneration (SCD) including sporadic olivopontocerebellar atrophy and familial cortical cerebellar atrophy. In the cerebellum, CaBP immunoreactivity was seen exclusively in the Purkinje cell in both SCD and control groups. However, the number of CaBP-immunoreactive Purkinje cells was significantly reduced in SCD. CaBP immunohistochemistry also disclosed abnormal morphological changes of Purkinje cells, which was not visualized on conventional strains or not clearly demonstrated on immunohistochemistry for neurofilaments. Moreover, reduced CaBP immunoreactivity was observed even in some remaining Purkinje cells of SCD suggesting that loss of CaBP precedes neuronal loss of Purkinje cell. We conclude that CaBP is a useful marker for Purkinje cell degeneration, and that reduced CaBP expression might have some association with the mechanism of the Purkinje cell degeneration in SCD.

Aged↗
PubMed1995

Conserved delta-activity in reverse enantiomeric opioid peptide.

A reverse enantiomeric peptide has a reversed amino acid sequence with enantiomeric amino acid residues compared with its parent peptide. In most cases the random change of amino acid sequence or chirality might be expected to bring about significant changes in peptide activity. However, the reverse enantiomeric peptides of Leu-enkephalin and Tyr-D-Ala-Gly-Phe-D-Leu (DADLE) have shown affinity for the opioid delta-receptor, but not for mu- or kappa-receptors. This suggests that delta-opioid receptor recognition occurs primarily through interaction with the peptide side chains, since the native opioid peptide and its reverse enantiomer are able to have similar side-chain conformation.

Amino Acid Sequence↗
PubMed1995

Histopathology of rabbit eyes with silicone-fluorosilicone copolymer oil as six months internal retinal tamponade.

Silicone-fluorosilicone copolymer oil has low viscosity (175-185 cSt) and is heavier than water (density, 1.16 g cm-3). Short term retinal tolerance (within 2 months) of the silicone-fluorosilicone copolymer oil has been reported to be the same as that of currently used intraocular silicone oil. Ocular response of the purified silicone-fluorosilicone copolymer oil were examined clinically and histopathologically from 2.5 months to 6 months after vitreous cavity injection in rabbit phakic eyes, and compared the oil tolerance with that of purified silicone oil (0.97 g cm-3, 5000 cSt). The effects in anterior chamber also were examined within 4 weeks of the silicone-fluorosilicone copolymer oil injection in different rabbits. Silicone-fluorosilicone copolymer oil recovered from the vitreous cavity at 6 months was analysed for cholesterol and retinol content by high performance liquid chromatography. Because of its low viscosity, silicone-fluorosilicone copolymer oil was easy to inject and remove from the vitreous cavity with a 20-G needle. After the vitreous injection, discrete droplet formation by the silicone-fluorosilicone copolymer oil occurred more easily than by silicone oil. Medullary ray detachment was seen in a silicone oil-, and some silicone-fluorosilicone copolymer oil-injected eyes at 4-6 months. Histopathologically, after 3-6 months disappearance of outer plexiform layer and disorganization of the photoreceptor layer of silicone oil-, and silicone-fluorosilicone copolymer oil-injected eyes were seen in the superior and the inferior retina, respectively. Migration of the photoreceptor cell nuclei to the photoreceptor layer was found in the inferior retina of silicone-fluorosilicone copolymer oil-injected eyes at 5-6 months. Small droplets ingested by mononuclear cells were found in the vitreous cavity or preretina at 4-6 months in silicone-fluorosilicone copolymer oil-injected eyes. After the anterior chamber injection, silicone-fluorosilicone copolymer oil induced endothelial cell damage in the area where the oil contacted continuously. Retinol and cholesterol were identified in silicone-fluorosilicone copolymer oil removed from the vitreous cavity. Silicone-fluorosilicone copolymer oil may be useful as an intraoperative device in retinal detachment surgery and as a short term (up to about 2 months) retinal tamponade but we do not recommend it for long term retinal tamponade.

Animals↗
PubMed1995

Analysis of the chromosomal localization of the repetitive sequences (RPSs) in Candida albicans.

The location and organization of repetitive sequences, members of the RPS family, which are sequences specific to Candida albicans, were determined on each chromosome of C. albicans strain FC18. Using pulsed-field gel electrophoresis, we separated seven fractions from eight chromosomes. Each chromosome was cleaved by BamHI and XhoI to excise the RPSs, which were then detected by hybridization with an RPS probe. All chromosomes except chromosome 4 carried RPSs, and these RPSs were located within a limited region on each chromosome. From the digestion of each chromosome with SfiI and probing with the RPSs, we found that these recognition sites within the RPS region were conserved among all RPS-containing chromosomes. For further characterization of the RPSs, the locations and the boundary regions of the RPSs were examined on chromosome 6 of strain FC18 as a model chromosome. Using the restriction enzymes SfiI, SmaI, XhoI, BamHI, MluI and NruI, we constructed a semi-macro physical map of the RPSs and their boundary regions on this chromosome. We also determined which part of the RPS was adjacent to each boundary by using sub-fragments of RPS as probes. The physical configuration around the RPSs and their boundary regions are presented. The results obtained should be useful for future analysis of the function of these regions.

Blotting, Southern↗
PubMed1995

Solution structure of a human calcitonin analog elucidated by NMR and distance geometry calculations.

Three-dimensional structure of a human calcitonin analog (abbreviated as hCTa) in which the amino acids of the wild type are replaced at position 12, 16 and 19 by leucine residues and further at position 22 by a tyrosine residue was studied in TFE solution by 1H-NMR and distance geometry calculations. This analog has a 15-20 times activity as compared with the wild type. The amino acid replacements resulted in formation of an amphiphilic alpha-helix in the region between the residues 4-20. The overall three-dimensional structure is similar to that of the wild type. The conformational feature of hCTa with a hydrophobic face composed with a Met and four Leu residues may be related to its higher hypocalcemic potency.

Amino Acid Sequence↗
PubMed1995

Soluble expression of a synthetic gene for human translation initiation factor 4E in Escherichia coli.

In order to obtain the active form of recombinant human initiation factor (eIF) 4E effectively, an artificial synthetic gene was cloned into an expression vector (pMAL-p2) and the soluble expression was attempted in Escherichia coli under the control of a tac promoter. Two expression systems were finally constructed as a fusion protein with maltose-binding protein, which contain a recognition sequence for the site specific protease alpha-thrombin and factor Xa, respectively. Most of the fusion protein was induced as a soluble form. The soluble human eIF-4E digested from the fusion protein showed binding specificity for the m7GTP affinity column.

Amino Acid Sequence↗
PubMed1995

Binding specificity of mutagenic tryptophan pyrolysates for DNA conformation: spectroscopic and viscometric studies.

The compounds, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), are major potent mutacarcinogens isolated from tryptophan pyrolysate. In order to investigate their interaction with DNA and effects on DNA conformation, studies involving circular dichroism, fluorescence and absorption spectroscopy and viscometric titration were performed. The results show that (a) Trp-P-1 and Trp-P-2 are potent intercalators of DNA with nearly the same specificity for the A-T and G-C (alternative purine-pyrimidine) base sequences, (b) the interaction of Trp-P-1 with the B-form of DNA is biphasic so that stiffening of the B-DNA conformation occurs over the range r ([Trp-P-1]/[DNA]) = 0-2.5, followed by transformation of B to the non-B conformation at r > 2.5, (c) the transformation to the non-B structure is not observed for Trp-P-2, although stiffening of the B-DNA conformation similarly occurs, and (d) both Trp-P-1 and Trp-P-2 promote unwinding of the salt-induced Z-DNA to give the B-form. These data indicate that the noncovalent interaction of Trp-P with DNA is mainly dependent on the B-form conformation.

Base Sequence↗
PubMed1995

[Gene expression of human eukaryotic initiation factor-4E for protein synthesis and study of its recognition mechanism of mRNA cap structure].

Being stimulated by the insights from model studies that (i) the intimate combination of hydrogen-bonding pairing and aromatic stacking interactions is important for the specific binding of guanine base by peptide and (ii) the pi-pi stacking force of Trp is significantly strengthened by the guanine N7-methylation (m7G), this research project was started, because (a) the mRNA cap structure is characterized by the existence of m7G and (b) an eukaryotic initiation factor-4E (eIF-4E), a protein which specifically recognizes the mRNA cap structure and opens the protein biosynthesis, contains 8 Trp residues irrespective of its relatively low molecular weight of about 25 kDa. In order to prepare the sufficient amount of sample for carrying out the analysis of the recognition mechanism of mRNA cap structure by eIF-4E at the atomic level, firstly, the expression of human eIF-4E gene in Escherichia coli was attempted. An artificial gene encoding for human eIF-4E was chemically synthesized and succeeded in the expression with two different forms, i.e., as a fusion protein with human growth hormone and a direct expression of soluble protein. The isolation of eIF-4E and its purification procedure using the m7GTP affinity chromatography were accomplished. It was shown by spectroscopic methods that the recombinant eIF-4E exhibits essentially the same tertiary structure as the native one and the binding ability with mRNA cap analog was identical with each other. In order to analyze the functional amino acid residues which are essential for specific recognition of mRNA cap structure, next a series of eIF-4E mutants were prepared by the site-directed mutagenesis, and His37, His200, Trp102 and Glu103 were suggested to be important for binding of mRNA cap structure, as judged from comparison of the binding abilities of respective mutants with a m7GTP affinity column. Since the crystals of recombinant eIF-4E-m7GTP complex suitable for X-ray crystallography are now in preparation, the detailed interaction mode between them will be opened in near future.

Amino Acid Sequence↗
PubMed1995

A 3-deazauracil-resistant mutant of Bacillus subtilis with increased production of cytidine.

Bacillus subtilis No. 344 is a cytidine-producing mutant strain derived from wild type strain No. 122. When 3-deazauracil-resistant mutants were derived from strain No. 344, some of the mutants had higher productivities of cytidine. Among them, strain No. 428 accumulated 14.2 mg/ml cytidine in the culture. Cytidine 5'-triphosphate (CTP) synthetase from strain No. 428 changed to be free from feedback inhibition by CTP, compared with the enzyme from strain No. 344.

Bacillus subtilis↗
PubMed1995

Solution conformation of mu-selective dermorphin and delta-selective deltorphin-I in phospholipid micelles, studied by NMR spectroscopy and molecular dynamics simulations.

Complete proton resonance assignments of the naturally occurring mu-selective dermorphin (H-Tyr-D-Ala-Phe-Gly-Tyr-Pro-Ser-NH2) and delta-selective deltorphin-I (H-Tyr-D-Ala-Phe-Asp-Val-Val-Gly-NH2) were carried out by two-dimensional 1H-NMR techniques to investigate the conformational features in the membrane-mimetic micelles of perdeuterated dodecylphosphocholine. Fifty possible three-dimensional structures for respective peptides were generated by means of distance geometry calculations, all of which satisfy the proton-proton distances derived from NOE measurements within the allowable range, and 25 of them were subjected to the molecular dynamics simulations for 10 ps, in which the NOE distances were included as the energetic constraints. Although conformers simulated for dermorphin showed relatively large conformational variations because of the limited NOE data, most of them were characterized as an entirely folded structure bent at the Gly4 residue, where each of the N- and C-terminal tetrapeptides took an extended conformation. On the other hand, most conformations of deltorphin-I showed the common feature that the N-terminal Tyr-D-Ala-Phe-Asp and C-terminal Val-Val-Gly-NH2 sequences took respective folded conformations, and these were almost at right angles on the border of the Asp-Val sequence. These conformational characteristics are discussed in terms of the possible relationship with the mu/delta-opioid receptor selectivity.

Amino Acid Sequence↗
PubMed1995

Retinoic acid in silicone and silicone-fluorosilicone copolymer oils in a rabbit model of proliferative vitreoretinopathy.

PURPOSE: The authors evaluated the effect of retinoic acid (RA) in silicone oil (SiO) and in silicone-fluorosilicone (SiFO) copolymer oil in a new rabbit model of proliferative vitreoretinopathy (PVR). METHODS: To create the PVR model, three groups of rabbits were administered vitreous injections of approximately 100,000 homologous fibroblasts, 75,000 platelet-rich plasma (PRP), and fibroblasts + PRP, respectively. These rabbits were followed up ophthalmoscopically and histopathologically for as long as 2 months. Five additional groups of rabbits underwent gas-compression vitrectomy in one eye. Four days later, group 1 was administered intravitreous RA in SiFO (9 micrograms/ml) with approximately 150,000 fibroblasts and 70,000 PRP. Group 3 was administered the same amount of fibroblasts and PRP as group 1 with RA in SiO (9 micrograms/ml). Groups 2, 4, and 5 were administered the same amount of fibroblasts and PRP as groups 1 and 3 with 1 ml of SiFO, SiO, or balanced salt solution only, respectively. To evaluate RA toxicity, RA was injected in SiO (15 and 20 micrograms/ml) and RA in SiFO (10 micrograms/ml). RESULTS: All eyes that were administered fibroblasts or PRP developed vitreous membranes, but those with PRP alone did not develop proliferative changes or retinal detachment; fibroblasts alone produced proliferative changes and retinal detachment after 2 to 3 weeks; fibroblasts + PRP produced similar changes within 3 days of injection. Retinoic acid (15 micrograms/ml) in SiO and RA (10 micrograms/ml) in SiFO was well tolerated. Retinal atrophic changes were found in eyes with 20 micrograms/ml RA in SiO. The retinal detachment rate was lower (P < 0.05) in the eyes that were administered fibroblasts + PRP and RA than in the controls. Significant differences were found in the degrees of PVR among the groups. CONCLUSIONS: RA could be useful in PVR treated with SiO or for eyes treated intraoperatively with heavier-than-water SiFO when it is used as a short-term retinal tamponade.

Animals↗
PubMed1994

Structure of ascidiacyclamide as the ethanol water solvate, a cytotoxic cyclic peptide from Ascidian.

The X-ray crystal structure determination of the C2H5OH.H2O solvate of ascidiacyclamide (C36H52N8O6S2), a cytotoxic cyclic peptide from marine tunicate Ascidian, revealed a C2-symmetric saddle-shaped rectangular conformation of the molecule. The water and ethanol molecules are located on the crystallographic diad axis and are held by hydrogen bonds and van der Waals contacts with the polar ring N atoms and nonpolar D-Val side-chain atoms, respectively. The molecular conformation and the interaction with solvent molecules are nearly the same as those of the compound with C2H5OH.2H2O [Ishida, In, Doi, Inoue, Hamada & Shioiri (1992). Biopolymers, 32, 131-143].

Animals↗
PubMed1994

DNA-binding characterization of a novel anti-tumour benzo[a]phenazine derivative NC-182: spectroscopic and viscometric studies.

NC-182 is a novel anti-tumour compound having a benzo[a]phenazine ring. Fluorescence, absorption and c.d. spectroscopy, as well as viscometric titrations, were systematically performed to investigate the interaction mode of this drug with DNA and its effect on DNA conformation, based on comparative measurements with distamycin (DNA minor-groove binder) and daunomycin (DNA-base intercalator). NC-182 was found to be a potent intercalator of DNA, especially the B-form DNA, although no specificity was observed against the base-pair. The binding of NC-182 to B-DNA behaves biphasically, depending on the molar ratio (r) of drug to DNA: NC-182 acts to render the B-form structure rigid at relatively low r value and to promote the transformation of B- to non-B forms at high r values. It was also shown that NC-182 promotes the unwinding of Z-form DNA to B-form. Viscometric, u.v. 'melting' and c.d. experiments further showed that (1) the DNA duplex structure is thermally stabilized by intercalation with NC-182 and (2) the intercalation of NC-182 into a poly(dA).2poly(dT) DNA structure thermally stabilizes the triplex structure, resulting in a melting point close to that of the duplex structure; the melting curves of triplex and duplex structures coincide at r > 0.06. These observations make a significant contribution to our understanding of the biological properties of this novel benzo[a]phenazine derivative, a new anti-tumour tumour agent against multidrug-resistant and sensitive tumours.

Antineoplastic Agents↗
PubMed1994

Conformational analysis of elcatonin in solution.

The conformational characteristics of elcatonin, an analogue of eel calcitonin having a disulfide bond Cys1-Cys7 replaced by an ethylene linkage between residues 1 and 7, have been analyzed in aqueous trifluoroethanol solutions. Circular dichroic spectra of elcatonin and eel calcitonin itself reveal the presence of alpha-helices at trifluoroethanol concentrations above 15%. The spectral changes caused by the trifluoroethanol content of the solutions are interesting. An isosbetic point is detected for eel calcitonin indicating that a conformational transition occurs between two states, namely alpha-helical and random coil states. On the other hand, the CD curves of elcatonin at less than 15% trifluoroethanol deviate from the isosbetic point while those at higher concentration are similar to those of eCT. This can probably be attributed to the third element of the ordered structure of elcatonin which is formed in 15% trifluoroethanol. The solution conformation of elcatonin in a mixture of 60% water and 40% trifluoroethanol has been determined by the combined use of 1H-NMR spectroscopy and distance geometry calculations. The conformation is characterized by an amphiphilic alpha-helix between Thr6 and Thr21, which extends into the constrained cyclic portion of the molecule to Thr6. The third structural element of elcatonin found in the CD analysis is detected by some turn structures in the region between residue 1 and Ser5 in the calculated structure.

Amino Acid Sequence↗
PubMed1994

Nucleotide sequence of the structural gene for the penicillin-binding protein 2 of Staphylococcus aureus and the presence of a homologous gene in other staphylococci.

The structural gene for penicillin-binding protein 2 (PBP2) of Staphylococcus aureus was cloned and sequenced. The nucleotide sequence of the 2,458-bp chromosomal fragment was determined, and the 2,148-bp coding region for PBP2 was identified. Determination of ten N-terminal amino acids of the PBP2 protein indicated that N-terminal methionine had been removed from the primary translational product. Thus, PBP2 is comprised of 715 amino acids with a molecular mass of 79,147. Nucleotide sequences having some homology with the PBP2 gene and proteins cross-reactive with anti-PBP2 antibody were detected in some other species of staphylococci by polymerase chain reaction and Western blot analysis, respectively.

Amino Acid Sequence↗
PubMed1994

A case of uveitis associated with idiopathic retroperitoneal fibrosis.

We treated a patient with nongranulomatous panuveitis associated with idiopathic retroperitoneal fibrosis who had symptoms of abdominal pain, severe fever, leg edema, and blurred vision. A high C-reactive protein level, a high erythrocyte sedimentation rate, anemia, and abnormalities of the immune system were seen. Retrograde pyelography disclosed persistent ureteral obstruction. A computed tomographic scan and magnetic resonance imaging showed a dense plaque of fibrous tissue around the kidney and hydronephrosis. Biopsy disclosed fibrosis infiltrated with lymphocytes, plasma cells, histiocytes, and eosinophils. A moderate degree of cellular inflammation in the anterior chamber and cystoid macular edema were present at ocular examination. We believe idiopathic retroperitoneal fibrosis to be a new cause of uveitis of suspected autoimmune origin.

Blood Sedimentation↗