Highly regioselective methylation of inosine nucleotide: an efficient synthesis of 7-methylinosine nucleotide

Abstract

A facile, straightforward, reliable, and an efficient chemical synthesis of inosine nucleotides such as 7-methylinosine 5′-O-monophosphate, 7-methylinosine 5′-O-diphosphate, and 7-methylinosine 5′-O-triphosphate, starting from the corresponding inosine nucleotide is delineated. The present methylation reaction of inosine nucleotide utilizes dimethyl sulfate as a methylating agent and water as a solvent at room temperature. It is noteworthy that the present methylation reaction proceeds smoothly under aqueous conditions that is highly regioselective to afford exclusive 7-methylinosine nucleotide in good yields with high purity (>99.5%).     

Crystal structure of the 270 kDa homotetrameric lignin-degrading enzyme pyranose 2-oxidase

Pyranose 2-oxidase (P2Ox) is a 270 kDa homotetramer localized preferentially in the hyphal periplasmic space of lignocellulolytic fungi and has a proposed role in lignocellulose degradation to produce the essential co-substrate, hydrogen peroxide, for lignin peroxidases. P2Ox oxidizes D-glucose and other aldopyranoses regioselectively at C2 to the corresponding 2-keto sugars; however, for some substrates, the enzyme also displays specificity for oxidation at C3. The crystal structure of P2Ox from Trametes multicolor has been determined using single anomalous dispersion with mercury as anomalous scatterer. The model was refined at 1.8A resolution to R and Rfree values of 0.134 and 0.171, respectively. The overall fold of the P2Ox subunit resembles that of members of the glucose-methanol-choline family of long-chain oxidoreductases, featuring a flavin-binding Rossmann domain of class alpha/beta and a substrate-binding subdomain with a six-stranded central beta sheet and three alpha helices. The homotetramer buries a large internal cavity of roughly 15,000 A3, from which the four active sites are accessible. Four solvent channels lead from the surface into the cavity through which substrate must enter before accessing the active site. The present structure shows an acetate molecule bound in the active site with the carboxylate group positioned immediately below the flavin N5 atom, and with one carboxylate oxygen atom interacting with the catalytic residues His548 and Asn593. The entrance to the active site is blocked by a loop (residues 452 to 461) with excellent electron density but elevated temperature factors. We predict that this loop is dynamic and opens to allow substrate entry and exit. In silico docking of D-glucose in the P2Ox active site shows that with the active-site loop in the closed conformation, monosaccharides cannot be accommodated; however, after removing the loop from the model, a tentative set of protein-substrate interactions for beta-D-glucose have been outlined.     

区域选择性改性的纤维素和壳聚糖衍生物及其血液相容性

通过对纤维素和壳聚糖的区域选择性改性,将内皮细胞表面硫酸乙酰肝素（ES—HS）分子结构中对其血液相容性有重要影响的官能团引入纤维素和壳聚糖的分子结构中,并将其通过离子键固定在部分阳离子化的纤维素膜上,以期模拟ES—HS的血液相容性。血小板吸附结果表明,6位改性的纤维素衍生物的吸附程度较高。在五种壳聚糖衍生物中,2位的NS03／NAc为6／4的衍生物表现出最低的血小板吸附。当保持壳聚糖2位的NS03／NAc值不变时,对6位进行完全磺酸酯化,也可有效减少血小板的吸附。     

Synthesis of regioselectively substituted cellulose derivatives and applications in chiral chromatography

Various cellulose-2,3-bis-arylcarbamate-6-O-arylesters and cellulose-2,3-bis-arylester-6-O-arylcarbamates, designed to test the possible combined effects of the known tris-arylcarbamate and tris-arylester classes, were synthesized with high regioselectivity at O-C(6), and their use as CSP s in liquid chromatography for enantiomeric separations was investigated. The separations obtained with the synthesized CSP s were compared to the separations achieved on a self-packed reference column, consisting of cellulose-tris-(3,5-dimethylphenyl-carbamate) as CSP standard. Among the synthesized, regioselectively substituted cellulose derivatives, 2,3-bis-O-(3,5-dimethylphenylcarbamate)-6-O-benzoate-cellulose and 2,3-bis-O-(benzoate)-6-O-(3,5-dichlorophenylcarbamate)-cellulose gave the best CSP s for the separation of the test racemates. CSP s from regioselectively substituted cellulose derivatives seem to exhibit higher selectivities than cellulose-tris-(3,5-dimethylphenylcarbamate) for certain classes of racemic compounds. Chirality 10:294–306, 1998. © 1998 Wiley-Liss, Inc.     

天然多羟基化合物在酶催化下的选择性酯化

综述了近十几年来几种天然多羟基化合物包括皂草苷类、黄酮苷类、生物碱类、甾醇类、倍半萜(烯)化合物、熊果苷和槐糖脂在脂肪酶和枯草杆菌蛋白酶催化下的选择性酯化。     

Regioselective oxidative phenol couplings of 2,3′,4,6-tetrahydroxybenzophenone in cell cultures of Centaurium erythraea RAFN and Hypericum androsaemum L.

A crucial step in plant xanthone biosynthesis is the cyclization of an intermediate benzophenone to a xanthone. In cultured cells of Centaurium erythraea RAFN, 2,3′,4,6-tetrahydroxybenzophenone (THBP) was shown to be intramolecularly coupled to 1,3,5-trihydroxyxanthone, whereas in cell cultures of Hypericum androsaemum L. it was coupled to form the isomeric 1,3,7-trihydroxyxanthone. These regioselective cyclizations that occur ortho and para, respectively, to the 3′-hydroxy group of the benzophenone depend on cytochrome P ₄₅₀, as shown by the effectiveness of established P ₄₅₀ inhibitors and blue-light-reversible carbon monoxide inhibition. Furthermore, the reactions absolutely require NADPH and O₂. The underlying reaction mechanism is probably an oxidative phenol coupling that is catalyzed regioselectively by xanthone synthases. These enzymes are proposed to be cytochrome P ₄₅₀ oxidases. The intramolecular cyclizations of THBP to 1,3,5- and 1,3,7-trihydroxyxanthones catalyzed by the two xanthone synthases represent an important branch point in the plant xanthone biosynthetic pathway. Received: 24 March 1997 / Accepted: 28 May 1997     

Syntheses and biological activities of deoxy-d-allose fatty acid ester analogs

We describe the syntheses of three different deoxy-D-allose analogs [2-deoxy-D-allose (2-DOAll), 1,2-dideoxy-D-allose (1,2-DOAll), and 1,2-didehydro-1,2-dideoxy-D-allose (1,2-DHAll)] and their fatty acid esters via regioselective lipase-catalyzed transesterification. Among them, 2-DOAll and its decanoate (2-DOAll-C10) showed higher inhibitory activity on plant growth, which is similar to D-allose (All) and its decanoate (All-C10). Bioassay results of deoxy-All-C10 on four plant species suggest that the hydroxy group at the C-1 position might be important showing growth inhibitory activity. In addition, co-addition of gibberellin (GA₃) with 1,2-DHAll-C10 and 2-DOAll-C10 recovered plant growth, suggesting that they might mainly inhibit biosynthesis of gibberellin.     

Regioselective enzymatic acylation of multi-hydroxyl compounds in organic synthesis

With current developments in enzyme-catalyzed reactions and techniques available for rational redesign of natural biocatalysts, the enzymatic biosynthesis can become one of the most valuable synthetic methods. Enzymatic regioselective catalysis in organic media has played a key role in pursuing asymmetric synthesis for active chiral compounds. Here, we shortly describe some historical issues of the rapidly growing area, enzymatic catalysis in synthetic organic chemistry and then review researches that have been carried out in the regioselective enzymatic catalysis for the past two decades. An application of this technology to the modification of some potential target drug compound will be also presented.     

Isolation and Structure Elucidation of Growth Inhibitors for Silkworm Larvae from Avocado Leaves

A cadmium and zinc-binding protein similar to metallothionein has been isolated from Tetrahymena pyriformis exposed to cadmium chloride. This protein contained 32.4% half-cystine, 23.7% acidic amino acids and 10.1% lysine. The amino acid composition was similar to that of copper-thionein of yeast. The metal-binding protein of Tetrahymena pyriformis contained 3.7 g atom cadmium, 0.7 g atom zinc, and O.l g atom copper per mol, and shpwed the spectral characteristics of cadmium-thionein, i.e., a broad shoulder at 250 nm and low residual absorption at 280 nm. The molecular weight of this protein was determined to be 11,000 by gel filtration in 6 M guanidine hydrochloride, although it moved like a protein with a molecular weight of 30,000 on ordinary gel filtration.     

Purification and Characterization of an Intracellular α-D-Xylosidase II from Aspergillus flavus MO-5

α-D-Xylosidase II activity from Aspergillus flavus MO-5 was increased roughly 5- to 10-fold by use of xylose instead of methyl α-D-xylopyranoside (α-MX) as a carbon source.

The enzyme was purified to an electrophoretically pure state by successive chromatography on Q-Sepharose, Phenyl Superose, PL-SAX, and TSK-gel G3000SWXL. The purified enzyme hydrolyzed isoprimeverose [α-D-xylopyranosyl-(1→6)-D-glucopyranose] and p-nitrophenyl α-D-xylopyranoside (α-p-NPX), but not α-MX or xyloglucan oligosaccharide. The apparent K_m and V_max of the enzyme for α-p-NPX and isoprimeverose were 0.97 mM and 28.0 µmol/min/mg protein, and 47.62 mM and 2.0 µmol/min/mg protein, respectively. This enzyme had an apparent molecular weight of 67,000 by SDS-polyacrylamide gel electrophoresis and 180,000 by gel filtration chromatography (TSK-gel G3000SWXL).

The enzyme showed the highest activity at pH 6.0 and 40°C, and was stable in the pH range from 6.0 to 7.0 and at the temperatures up to 40°C. The activity was inhibited by Cu²⁺, Zn²⁺, Hg²⁺, p-CMB, SDS, Fe³⁺, and N-ethylmaleimide.

This enzyme had nothing in common with α-D-xylosidase I and four α-D-xylosidases reported already.