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Minoru Isobe

Publications and source records attributed to Minoru Isobe.

At least 37 records · Page 2Linked to original sources
PubMed2005

Allyl cyanate-to-isocyanate rearrangement for the synthesis of quaternary stereocenter with nitrogen substituent.

The stereochemistry and efficiency of an allyl cyanate-to-isocyanate rearrangement for the construction of quaternary stereocenter with nitrogen substituent was investigated by the synthesis of (R)-alpha-methylphenylalanine. The rearrangement was found to be stereospecific, and the chirality of allyl carbamate was transferred to that of the quaternary carbon bearing isocyanate group. These results establish that an allyl cyanate-to-isocyanate rearrangement is a useful method for the synthesis of natural products, that contain the quaternary carbon with nitrogen substituent.

Allyl Compounds↗
PubMed2004

Aluminum as a specific inhibitor of plant TPC1 Ca2+ channels.

In plant cells, Al ion plays dual roles as an inducer and an inhibitor of Ca(2+) influx depending on the concentration. Here, the effects of Al on Ca(2+) signaling were assessed in tobacco BY-2 cells expressing aequorin and a putative plant Ca(2+) channel from Arabidopsis thaliana, AtTPC1 (two-pore channel 1). In wild-type cells (expressing only aequorin), Al treatment induced the generation of superoxide, and Ca(2+) influx was secondarily induced by superoxide. Higher Al concentrations inhibited the Al-stimulated and superoxide-mediated Ca(2+) influx, indicating that Ca(2+) channels responsive to reactive oxygen species (ROS) are blocked by high concentration of Al. H(2)O(2)-induced Ca(2+) influx was also inhibited by Al. Thus, inhibitory action of Al against ROS-induced Ca(2+) influx was confirmed. Similarly, known Ca(2+) channel blockers such as ions of La and Gd inhibited the H(2)O(2)-induced Ca(2+) influx. While La also inhibited the hypoosmotically induced Ca(2+) influx, Al showed no inhibitory effect against the hypoosmotic Ca(2+) influx. The effects of Al and La on Ca(2+) influx were also tested in the cell line overexpressing AtTPC1 and the cell line AtTPC1-dependently cosuppressing the endogenous TPC1 equivalents. Notably, responsiveness to H(2)O(2) was lost in the cosuppression cell line, thus TPC1 channels are required for ROS-responsive Ca(2+) influx. Data also suggested that hypoosmotic shock induces TPC1-independent Ca(2+) influx and Al shows no inhibitory action against the TPC1-independent event. In addition, AtTPC1 overexpression resulted in a marked increase in Al-sensitive Ca(2+) influx, indicating that TPC1 channels participate in osmotic Ca(2+) influx only when overexpressed. We concluded that members of TPC1 channel family are the only ROS-responsive Ca(2+) channels and are the possible targets of Al-dependent inhibition.

Aequorin↗
PubMed2004

Total synthesis of (+)-blasticidin s.

The first total synthesis of the peptidyl nucleoside antibiotic, blasticidin S (1), has been achieved by the coupling reaction of cytosinine (3) and blastidic acid (2). A key step in the synthesis of cytosinine (3) is the sigmatropic rearrangement of allyl cyanate 24; this reaction provided efficient and stereoselective access to 2,3-dideoxy-4-amino-D-hex-2-enopyranose (26 a). Further elaboration of 26 a gave methyl hex-2-enopyranouronate 29, and cytosine N-glycosylation of 31 using the Vorbrüggen conditions for the silyl Hilbert-Johnson reaction furnished the differentially protected cytosinine (32) in 11 steps from 2-acetoxy-D-glucal (14) (4.0 % overall yield). Synthesis of the Boc-protected blastidic acid 47 in nine steps starting from chiral carboxylic acid 35 (23 % overall yield) utilized Weinreb's protocol for the preparation of benzyl amide 38 and Fukuyama's protocol for the synthesis of the secondary amine 40. Assembly of the protected cytosinine (32) and blastidic acid (47) by the BOP method in the presence of HOBt, and finally elaboration to 1 by deprotection of the fully protected 54 established the total synthesis of blasticidin S (1).

Amino Acids↗
PubMed2004

Structure of pentasaccharide of glycopeptide from TIME-EA4, N-glycoprotein in silkworm diapause eggs.

The TIME-EA4, from silkworm diapause eggs of pure strain C108, Bombyx mori, has glycosylated chain as tetrasaccharide (Man(2)GlcNAc(2)) attaching to the Asn(22) of T3 peptide from tryptic digests. On the other hand, from Showa silkworm strain we additionally observed a pentasaccharide (Man(3)GlcNAc(2)) on T3 at the same linkage site. The linkage pattern of the 5-sugar chain was studied through Smith degradation combined with LC-MS and MS/MS analyses. These advanced methods led us to conclude that the pentasaccharide was branching as Man 1-->3(Man 1-->6)Man 1-->4GlcNAc 1-->4GlcNAc.

Animals↗
PubMed2004

Alpha-C-mannosyltryptophan is not recognized by conventional mannose-binding lectins.

Alpha-C-mannosyltryptophan (C-Man-Trp) is a novel, naturally occurring C-linked carbohydrate-protein linkage first found in 1994 from human ribonuclease 2. Since then, a number of C-Man-Trp residue have been found from several important proteins such as interleukin 12 beta, components of complement system, thrombospondin-1, and erythropoietin receptor, however, the biological functions have remained unknown even though its biosynthetic pathway has been revealed. In order to find a clue as to the biological functions, we examined the affinity of C-Man-Trp with conventional mannose lectin such as concanavarin A (Con A) and mannose-binding lectin (MBL). The affinity of C-Man-Trp with Con A, a typical mannose-binding lectin from plant was examined using a Con A-Sepharose column. Unlike p-nitrophenyl-alpha-O-Man, C-Man-Trp was not retained on the column. MBL-C, a major mannose-binding lectin purified from mouse serum, did not bind with N-biotinylated C-Man-Trp, judging from ELISA based assay. These results imply that C-Man-Trp may be recognized with the other specific proteins associated with its unknown biological functions.

Blotting, Western↗
PubMed2004

Stereocontrolled synthesis of 8,11-dideoxytetrodotoxin, an unnatural analogue of puffer fish toxin.

8,11-Dideoxytetrodotoxin, an unnatural tetrodotoxin analogue, was synthesized in a highly stereoselective manner from a common intermediate from our synthetic studies on tetrodotoxin. The key features in the synthesis were as follows: neighboring group participation of a trichloroacetamide to allow regioselective and stereoselective hydroxylation, protection of a delta-hydroxylactone as an ortho ester, and guanidine installation through the use of Boc-protected isothiourea. Global deprotection of the fully protected intermediate under acidic conditions gave 8,11-dideoxytetrodotoxin, which exhibited very weak biological activities.

Animals↗
PubMed2004

Differentiation of sulfate and phosphate by H/D exchange mass spectrometry: application to isoflavone.

Often phosphorylation or sulfation is an important step which occurs in the signal transduction and cascade of metabolic pathways. Some natural products and metabolites contain one or more sulfate or phosphate groups. Isoflavone sulfate has been identified from high-resolution mass spectrometry (HRMS) and enzymatic digestion by sulfatase. We previously reported the new water-soluble isoflavone analogs, daidzein 7-O-phosphate and genistein 7-O-phosphate, which were surprisingly hydrolyzed by sulfatase. In this previous study, we could not determine the phosphate from the results of HRMS and enzymatic digestion, that is, HRMS and enzymatic digestion did not provide clear evidence. In this case, we drew conclusions from NMR analysis. HRMS has been ineffective with a regular fast atom bombardment (FAB) mass spectrometer to distinguish between phosphate and sulfate since the mass difference is only 0.009 mass units. There was, however, no conventional method of microanalysis to distinguish phosphate from sulfate owing to the same nominal mass. It is still very difficult to determine by negative FABMS [--O--P(==O)(OH)(2)] = 80 and [--O--S(==O)(2)OH] = 80. In this paper, we report a method to distinguish between these groups by using a popular low-resolution mass instrument; thus, phosphate and sulfate were measured by H/D exchange mass spectrometry at the picomole level to differentiate [--O--P(==O)(OD)(2)] = 82 and [--O--S(==O)(2)OD] = 81, respectively. This method is applicable not only to the isoflavone, but also to other phospho and sulfo compounds.

Genistein↗
PubMed2004

Demarcation of diapause development by cold and its relation to time-interval activation of TIME-ATPase in eggs of the silkworm, Bombyx mori.

We investigated the mode of action of winter cold in the termination of diapause by investigating Time-Interval-Measuring Enzyme (TIME). First, we determined the period of cold required for the completion of diapause development. Synchronously developing egg batches of a pure strain (C108 Bombyx mori silkworm) were used to minimize variations in hatching time. Hatching occurred with only 18 days chilling at 5 degrees C when the incubation at 25 degrees C after the chilling was elongated. The 18-day period was much shorter than we expected; diapause in B. mori is known to terminate completely with about 100 days of chilling. Even in such a short period of chilling, no sporadic hatching occurred. Moreover, we determined that a temperature-insensitive stage, which we called "Neboke", followed the short cold-requiring stage. Thus, the stage of diapause development was demarcated from other stages of diapause. While the length of diapause development was elongated when chilling was delayed after oviposition, the Neboke stage length was invariant. Cold evidently exerts its effect only on diapause development. When TIME was purified from eggs and chilled in test tubes, a transitory burst of its ATPase activity occurred at a time equivalent to shortly before the completion of diapause development; this was an interval-timer activation. The mechanism by which cold activates TIME to measure the time interval may help explain in biochemical terms the insect's adaptation to its seasonal environments.

Acclimatization↗
PubMed2004

Mass spectrometric assignment of Smith degradation glycopeptides derived from ribonuclease B.

We established a method to determine the glycosyl linkage structure by a combination of Smith degradation and liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry (LC-ESI-Q-TOF-MS) and tandem MS (MS/MS). To assign the sugar linkage of N-glycoprotein, we employed a typical ribonuclease B containing oligosaccharides (Man5-9GlcNAc2). Tryptic digestion of ribonuclease B provided a mixture of high-mannose glycopeptides consisting of the four amino acids, Asn34-Leu-Thr-Lys37 (NLTK, T6). The mixture of glycopeptides was separated by high-performance liquid chromatography (HPLC) in a reversed phase column and was characterized by ESI-Q-TOF-MS and MS/MS. Comparison of the data with and without Smith degradation allowed us to make reasonable assignments to support such linkage patterns as (1-->2), (1-->3), (1-->6) and their multiples. These assignments were limited to six mannoses or lower due to the unstable nature of the higher derivatives. This method should be applicable to determine the linkage pattern of an unknown glycoprotein in about a 6-microgram amount.

Amino Acids↗
PubMed2003

Complexation of cyclic dodecadepsipeptide, cereulide with ammonium salts.

Cereulide is a principal toxin causing emetic syndrome which is produced by Bacillus cereus and has been known as potassium selective ionophore. This paper deals with its complexation with inorganic and organic ammonium ions to assign the higher structures similar to the complex with potassium ion by means of NMR and ESI-MS spectroscopy. Of particular interest, the detectable ions are not only at m/z 1191.8 as K(+) complex but also (or sometimes exclusively) at m/z 1170.8 as NH(4)(+) complex in its LC-MS analyses depending upon the conditions. This difference is due to the sample preparation and measurement condition.

Depsipeptides↗
PubMed2003

First asymmetric total synthesis of tetrodotoxin.

Tetrodotoxin, a toxic principle of puffer fish poisoning, is one of the most famous marine natural products because of the complex structure having many functional groups and its potent biological activity leading to death. Since the structure elucidation in 1964, this toxin has been recognized as a formidable target molecule for total synthesis. We have recently achieved the first asymmetric total synthesis from 2-acetoxy-tri-O-acetyl-d-glucal as a chiral starting material. The highly hydroxylated cyclohexane ring was constructed by Claisen rearrangement and regioselective hydroxylations of an acetone moiety and an intramolecular directed aldol condensation of the precursor having methyl ketone with dihydroxyacetone, which was synthesized through Sonogashira coupling. Installation of nitrogen functionality was unsuccessful through an attempted Overman rearrangement. We, therefore, employed a new intramolecular conjugate addition strategy between the carbamate and unsaturated ester groups. The alpha-hydroxyl lactone moiety was synthesized through an intramolecular epoxide opening by the Z-enolate of aldehyde, which was followed by oxidation-reduction of the resulting cyclic vinyl ether. The lactone was then converted to a protected ortho ester, and then gunanidinylation was followed by cleavage of the 1,2-glycol to give the fully protected tetrodotoxin. Selection of the protective groups has finally led us to accomplish the total synthesis of tetrodotoxin in an enantiomerically pure form. All the stereogenic centers were controlled with high selectivity, and the hydroxyl groups were differently protected to discriminate for the future analogue synthesis of a bioorganic program. The synthetic tetrodotoxin was purified by ion exchange chromatography and characterized to be identical with the natural compound.

Cyclohexanes↗
PubMed2003

Convergent synthesis of the E'FGH ring fragment of ciguatoxin 1B via an acetylene cobalt complex strategy.

A convergent synthesis of the E'FGH ring fragment 28 of ciguatoxin 1B, a principal toxin causing widespread seafood poisonings "ciguatera", has been accomplished through (i) coupling between the E' ring-acetylide 9 and the H ring-aldehyde 20, (ii) stereoselective F ring cyclization via an acetylene cobalt complex, (iii) conversion to a carbonyl function under high-pressure hydrogenation, and (iv) reductive hydroxyketone cyclization to construct the G ring. In the (1)H NMR analysis of 28 at room temperature, a considerable broadening phenomenon was observed due to the slow conformational changes of the FG ring, as reported for natural ciguatoxin 1B. When measured in pyridine at -20 degrees C, the spectra of 28 exhibited a 3.5:1 mixture of two conformational isomers (UP and DOWN conformers).

Journal Article↗
PubMed2003

Biological activity of 8,11-dideoxytetrodotoxin: lethality to mice and the inhibitory activity to cytotoxicity of ouabain and veratridine in mouse neuroblastoma cells, Neuro-2a.

Contribution of the C-8 hydroxyl group of tetrodotoxin to its sodium channel blocking activity has never been clearly evaluated. Isobe et al. recently synthesized 8,11-dideoxytetrodotoxin, the first 8-deoxy analog of tetrodotoxin. In this study, the biological activity of this compound was investigated to compare with that of 11-deoxytetrodotoxin. Intraperitoneal injection of 8,11-dideoxytetrodotoxin at the level of 700 microg/kg did not kill a mouse (n=2), indicating that the lethal dose of this compound was more than 70 and 10 folds larger than LD(50) of tetrodotoxin and 11-deoxytetrodotoxin, respectively. The inhibitory activity of 8,11-dideoxytetrodotoxin to cytotoxicity of ouabain and veratridine in mouse neuroblastoma cells (Neuro-2a) was also examined. The ED(50) for 8,11-dideoxytetrodotoxin was estimated to be 9.3+/-3.3 microM (n=3), approximately 2000 and 34 folds larger than those of tetrodotoxin (4.6+/-0.70 nM, n=3) and 11-deoxytetrodotoxin (270+/-74 nM, n=4), respectively. These data suggest that the C-8 hydroxyl group of tetrodotoxin is also important for its activity, as well as all the other hydroxyl groups.

Animals↗
PubMed2002

Stereocontrolled synthesis of 8,11-dideoxytetrodotoxin, unnatural analogue of puffer fish toxin.

[structure: see text] 8,11-Dideoxytetrodotoxin, an unnatural tetrodotoxin analogue, was synthesized in a highly stereoselective manner from a common intermediate in our synthetic studies on tetrodotoxin. The synthesis features neighboring group participation of trichloroacetamide for stereoselective hydroxylation, protection of ortho ester, and guanidine installation with Boc-protected isothiourea.

Stereoisomerism↗
PubMed2002

Asymmetric total synthesis of 11-deoxytetrodotoxin, a naturally occurring congener.

Tetrodotoxin, a toxic principle of puffer fish poisoning, is a specific blocker of sodium channel. Despite many synthetic efforts since the structure elucidation in 1964, the only total synthesis of the racemic tetrodotoxin has been reported by Kishi and co-workers. In the course of our studies directed toward the total synthesis to analyze biologically interesting issues associated with tetrodotoxin, we accomplished a highly stereocontrolled synthesis of (-)-5,11-dideoxytetrodotoxin in 1999. Based on the synthesis, we describe herein the first total synthesis of 11-deoxytetrodotoxin, a naturally occurring analogue. The synthesis started from an allylic alcohol, the same intermediate for the synthesis of 5,11-dideoxytetrodotoxin. Epoxidation of the allylic alcohol was followed by isomerization with Ti(i-PrO)(4) to give an alpha-hydroxy allylic alcohol, in which the configurations of the two hydroxyl groups were inverted by oxidation and then a 2-step reduction. Further epoxidation of the allylic alcohol and ozonolysis of the remaining vinyl group gave an aldehyde, which reacted with magnesium acetylide to give a propargyl alcohol in a stereoselective manner. Oxidative cleavage of the acetylenic moiety with RuO(4) afforded a fully functionalized lactone for 11-deoxytetrodotoxin. Crucial guanidinylation was achieved from trichloroacetamide according to our own method to give acetyldibenzylguanidine. Finally, deprotection of benzyl groups, acetates, and acetal furnished 11-deoxytetrodotoxin.

Animals↗
PubMed2002

A novel photoprotein from oceanic squid (Symplectoteuthis oualaniensis) with sequence similarity to mammalian carbon-nitrogen hydrolase domains.

A 60-kDa photoprotein was selectively extracted from squid photogenic organ with 0.6 M KCl solution at pH 6 as luminescence-active forms. The photoprotein with fluorescence chromophore was eluted from size-exclusion HPLC mainly as oligomeric forms (about 200 kDa or more) with a trace amount of monomeric form of about 60 kDa. A limited tryptic digestion of the KCl-extract induced the cleavage into a 40-kDa fragment and a 16-kDa N-terminal fragment and the conversion to the monomeric form which still retained luminescence activity. Under UV light the 60-kDa protein and its 40-kDa fragment emitted fluorescence. Immunoblot analysis using specific antibody showed specific expression of the 60-kDa protein in the photogenic organ. Amino acid sequences of the 60-kDa photoprotein, its 40- and 16-kDa fragments, and six peptides from the Lys-C digest revealed no sequence similarity to known photoproteins but significant similarity to the carbon-nitrogen hydrolase domain found in mammalian biotinidase and vanin (pantetheinase).

Amidohydrolases↗