Succesful structural transformations of bioactive compounds into newer skeletal structures by replacing the substructure with others, the fedtures of which are not necessarily similar to but more or less drastically varied from the originial one, were proposed to be called being made "bioanalogously" instead of "bioisosterically". Precedents of the bioanalogous replacements of substructures composed of the amide, urea, and related components with others were explored. Anilides, N-phenylureas, and N-phenylcarbamates are bioanalogous as herbicides and topical antiseptics. The bioanalogy can be expanded to include substructures containing ester as well as ether components when local anesthetics are considered together. The polar hydrogen-bondinig groups such as (thio)urea, cyanoguanidine, and nitroethenediamine substructures found in histamine H₂-receptor antagonists are also bioanalogous in various other bioactive compound series. The open-chain amides and the corresponding "carbonylogously" ring-closured dicarboximides are bioanalogous in agrochemicals and antiandfogens as well as in CNS (central nervous system)-active agents. Very often, similarities in the substructural transformation patterns are observed in various bioanalogous series regardless of differences in the pharmacological category. The observations could be used to predict newer generation structures from an ultimate lead structure.

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The concentration debendence of the mechanical properties of starch pastes near the sol-gel transition point was analyzed by the scaling law derived from percolation theory. Since the storage modulus curves for a starch paste slowly decrease with decreasing frequency, the plateau region at a frequency of 10^-1 s^-1 is shown to represent the elastic modulus that characterizes the network structure. The percolation threshold can be taken as 1.5 wt% for corn starch and as 1.0 wt% for cassava starch. The storage modulus was scaled on C-C_c (concentration-critical concentration, the critical exponents being determined as 4.2 and 1.8 for corn starch and cassava starch, respectively. It is suggested from these results that the intermolecular bonding of corn starch was rigid and that of cassava starch was flexible.

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Okara Koji (OK) is the term for okara fermented by Aspergillus oryzae (a fungus for making Miso Koji). The amount of sterol excreted in the feces from rats fed with OK as the dietary fiber (DF) source was greater than that from rats fed with okara (OC, the insoluble residue of homogenized soybean) as the DF source. The digestive residue of OK's insoluble dietary fiber (IDF) was more porous than that of OC-IDF. The stronger sterol excretion with OK would have arisen from the complementary action of OK-IDF hemicellulose and the porosity of the OK-IDF digestive residue. Their complementary action would contribute to the cholesterol-lowering action of OK. The levels of raffinose and stachyose in the OK diet were lower than those in the OC diet. The intake and utilization of soluble dietary fiber and insoluble dietary fiber by the OK diet group were lower than those by the OC diet group. OK would not be a cause of meteorism. Iron absorption with the OK diet was greater than that with the OC diet, and there may have been little inhibition of iron absorption by phytic acid in the OK diet. These results suggest that OK would be a useful food material as a DF source.

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Of several organic nitrogen sources tested, corn steep liquor (CSL) was found to be the most suitable for cellulose production by Acetobacter xyliuum subsp. sucrofermentans BPR 2001. When lactate which was detected only in CSL, was added to culture media containing other nitrogen sources, cellulose production was stimulated to levels similar to that in CSL medium. Lactate was found to stimulate cell growth during the early stage of culture and shown to function by linking with the respiratory chain and generating energy for growth. Therefore, it was speculated that lactate acts as an accelerator driving the TCA cycle as well as an energy producer, resulting in high cellulose production and rapid cell growth. The effects of various amino acids were also investigated: L-methionine was found to be essential for high cellulose yields and stimulation of cell growth during the eaily stage of culture. On the basis of these observations, a defined synthetic medium, containing lactate and methionine, was formulated, with which cellulose production of 90% of that with the CSL medium was reached in jar fermentor cultures.

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Lecithin: cholesterol acyltransferase (LCAT) is an enzyme that is important in the cholesterol reverse-transport system by converting the surface cholesterol of high-density lipoprotein (HDL) to cholesterol ester inside HDL. We have investigated changes in plasma LCAT activity induced by oxidized low-density lipoprotein (ox-LDL). (i) LCAT activity was inhibited by the addition of even a small amount of ox-LDL, (ii) This inhibitory effect was dose-dependent and was related to the degree of oxidation of LDL, (iii) As this inhibitory effect was not prevented by antioxidants, it is suggested that it does not occur via oxidation of substrate HDL. These results suggest that cholesterol reverse-transport via HDL may be affected by ox-LDL and they are noteworthy in suggesting a new physiological function for ox-LDL.

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Basic fibroblast growth factor (bFGF)-like immunoreactivity was detected in extracts of Panax ginseng root by using a sensitive two-site enzyme immunoassay specific for human bFGF (hbFGF). In an investigation of the molecular properties of this bFGF-like molecule (bFGF-LI), the bFGF-LI and hbFGF were found to be equivalent with respect to antigenicity, molecular weight, isoelectric point, affinity for binding to heparin, and mitogenic activity toward BALB/c3T3 fibroblasts. The identification of this bFGF-LI molecule in Panax ginseng root helps to explain various activities of the traditional Chinese medicine ginseng.

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Herein the structures and peel contents of anthocyanins from 26 kinds of 11 species of edible legumes are reported. In the scarlet bean (Phaseolus coccineus) delphinidin 3-glucoside was found, and in the sultanipya bean (Phaseolus lunatus) peonidin 3-glucoside and peonidin 3-rutinoside. In the black turtle bean (Phaseolus vulgaris) delphinidin 3-glucoside. petunin, petunidin 3-glucoside, malvin, and malvidin 3-glucoside were assayed. We also succeeded in the isolation and structural analysis of an anthocyanin from the adzuki bean ( Vigna angularis) as a cyanin for the first time. The anthocyanin contents of black colored seed coats were very high at around 0.2% for peel, and these beans may therefor be expected to find applications as food colorants. In individual vigna species from different districts, consistency in the variety of anthocyanins was observed.

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Two novel methods to detect reducing sugar on a thin-layer chromatographic plate, using aqueous coloring reagents and a commercial microwave oven, were developed. After spraying the modified Somogyi reagent on the plate, irrigating the reducing sugars, and then heating in a commercial microwave oven for a few minutes, the modified Nelson reagent was sprayed on the plates. Reducing sugars were only apparent as blue spots. On the other hand, after spraying the bicinchoninate reagent on the plates and then heating in the microwave oven, the sugar spots became reddish-violet. These two new methods enabled 0.1μg of glucose per spot to be detected on a TLC plate. Non-reducing sugars (sucrose, trehalose, methyl α-D-glucoside, and transfer products containing non-reducing ends) were not detectable by these methods.

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Somatic embryogenesis that occurred directly from carrot epidermal cells was monitored by scanning electron microscopy. Epidermal cells peeled off from hypocotyl tissues were visible as compressed cells, which expanded to become tubular cells. Subsequently, tubular cells divided regularly and "horizontally"to form linear clusters of cells and then these clusters of cells divided "vertically" to begin development. Swelling structures were formed by irregular and oblique division at sites on the linear clusters of cells and these structures continued cell division to form small proembryos. Some cells at the surface of the swelling structures and proembryos expanded in every direction to cover the surface of each structure. These expanding cells appeared to form a surface layer and then cell layers piled up upon one another as the swelling proceeded to a globular-stage embryo via a small proembryo.

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In a culture filtrate of Trichosporon penicillatum B2, which is a γ-ray irradiation mutant induced from T. penicillatum SNO3, we found three kinds of pectin-releasing enzymes, protopectinases SE1, SE2, and SE3, that have endo-polygalacturonase activity. These enzymes were purified to homogeneity with cation-exchange and size exclusion chromatographies. The major PPase in the culture filtrate was PPase SE1, which accounted for 75% of total activities in the culture filtrate, and the two others were 0.15% (PPase SE2) and 0.007% (PPase SE3). Their molecular masses were approximdtely 41, 41 and 42 kDa on SDS-PAGE, respectively. They had similar enzymatic propertied but different PPase activity and pH- and thermo-stability. Antibody against PPase S, which is produced by strain SNO3, inhibited the activities of PPase SE1, SE2, and SE3. However PPase SE1 was completely inhibited by treatment with the anti-PPase S antibody, but the activities of PPases SE2 and SE3 remained at 20 and 50% of the original activity, respectively.

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A novel lectin (GN_LL) was isolated from the leaves of the Great Northern bean, Phaseolus vulgaris. GN_LL was purified by affinity chromatography on ovomucoid-Sepharose 4B. GN_LL had a molecular mass of 135 kDa on gel filtration and gave two bands on SDS-polyacrylamide gel electrophoresis (PAGE) (band A of 34.0 kDa and band B of 34.2 kDa). Binding assay of horseradish peroxidase (HRP)-glycoproteins to the bands electroblotted onto polyvinylidene difluoride (PVDF) membrane showed that both bands could bind to complex-type N-linked oligosaccharide chains in glycoproteins. The N-terminal amino acid sequences of both bands were identical through the 10 residues and identical to that of α-subunit of a pod lectin (pod-α-subunit) from the same bean. On the other hand, band B cross-reacted with monoclonal antibody against a seed lectin from the same bean, but band A did not.

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An N-carbamyl-L-amino acid amidohydrolase was purified from cells of Escherichia coli in which the gene for N-carbamyl-L-amino acid amidohydrolase of Pseudomonas sp. strain NS671 was expressed. The purified enzyme was homogeneous by the criterion of SDS-polyacrylamide gel electrophoresis. The results of gel filtration chromatography and SDS-polyacrylamide gel electrophoresis suggested that the enzyme was a dimeric protein with 45-kDa identical subunits. The enzyme required Mn²⁺ ion (above 1 mM) for the activity. The optimal pH and temperature were 7.5 and around 40°C, respectively, with N-carbamyl-L-methionine as the substrate. The enzyme activity was inhibited by ATP and was lost completely with p-chloromercuribenzoate (1 mM). The enzyme was strictly L-specific and showed a broad substrate specificity for N-carbamyl-L-α-amino acids.

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Liposomal suspensions of dilinoleoylphosphatidylcholine (DLPC) containing α-tocopherol (0.1mol%, based on DLPC) were oxidized at 37°C. The oxidation was initiated by a lipid-soluble or water-soluble free radical initiator, or by the addition of CuSO₄ and fructose. In all the oxidation systems, α-tocopherol suppressed the formation of DLPC hydroperoxides until all the α-tocopherol had been depleted. The oxidation products of α-tocopherol were 8a-alkyldioxy-α-tocopherones, 5, 6-epoxy-α-tocopherylquinone, 2, 3-epoxy-α-tocopherylquinone, and α-tocopherylquinone. The 8a-alkyldioxy-α-tocopherones were decomposed in the liposomes primarily by being hydrolyzed to produce α-tocopherylquinone. The results indicate that α-tocopherol can trap peroxyl radicals to form 8a-alkyldioxy-α-tocopherones which are hydrolyzed to α-tocopherylquinone in phospholipid bilayers. In another oxidation pathway, α-tocopherol may be oxidized by peroxyl radicals to form isomeric epoxy-α-tocopherylquinones.

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Der fI is a major mite proteinaceous allergen found in house dust. We produced recombinant Der fI (reDer fI) in insect cells using a baculovirus expression system. Based on the molecular mass and N-terminal amino acid (aa) sequence analysis, reDer fI was found to be comprised of a mixture of two proteins, each of which includes a pro-sequence of different length. The reDer fI had IgE-binding activity at only 20% of that of native Der fI. The removal of the pro-sequence in an acidic solution drastically increased IgE-binding activity to almost the same as that of native Der fI, showing that the presence of the pro-sequence is inhibitory to the IgE-binding activity of Der fI.

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A sulfur-oxidizing bacterium strain NB1-3 isolated from corroded concrete was a Gram negative, non-spore-forming, and rod-shaped bacterium (0.5-1.0×1.5-2.0 μm) with a polar flagellum. Strain NB1-3 had its optimum temperature and pH for growth at 30°C and 3.0-4.0, respectively. Strain NB1-3 had enzyme activities that oxidized elemental sulfur, thiosulfate, tetrathionate, and sulfide and the activity to incorporate ¹⁴CO₂ into the cells. The mean G + C content of the DNA was 52.9 mol%. These results indicate that strain NB1-3 is Thiobacillus thiooxidans. Since nickel has been known to protect concrete from corrosion, the effect of Ni on the growth of strain NB1-3 was studied. The cell growth on tiosulfate-, elemental sulfur-, or tetrathionate-medium was completely inhibited by 0.1% metal nickel or 5 mM NiSO₄. Both cellular activities of elemental sulfur oxidation and CO₂ incorporation were strongly inhibited by 5 mM NiSO₄. The amounts of Ni in cells with or without nickel treatment were 1.7 and 160.0 nmol/mg protein, respectively. These results indicate that nickel binds to strain NB1-3 cells and inhibits enzymes involved in sulfur oxidation of this bacterium, and as a result, inhibits cell growth.

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Amino acid residue 56 of the phosphatidylserine synthase of Bacillus subtilis was changed from Ser to Pro by using modified primers in PCR amplification of its structural gene, pss_BS. When an Escherichia coli mutant lacking its own phosphatidylserine synthase harbored a plasmid carring this allele, the Mn²⁺ -requiring Bacillus-type synthase activity, as assayed in vitro, was at least six-fold lower than that with the wild-type pss_BS gene and the cellular phosphatidyelethanolamine content was similarly lowered, indicating that the altered region of the enzyme is critically important for its activity. In contrast to the E. coli counterpart, amplification of the wild-type Bacillus enzyme increased both the relative and absolutecontents of phosphatidylethanolamine and impaired cell growth. However, amplification of the mutant enzyme of the same level was much less toxic, implying that E. coli cells are more sensitive to the unbalanced accumulation of phosphatidylethanolamine than that of the hydrophobic enzyme molecules. Possible roles of the conserved region of the enzyme in its activity and the wild-type phospholipid composition in the proper membrane function are discussed.

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An amylopullulanase (APase) from alkalophilic Bacillus sp. KSM-1378 hydrolyzes both α-1, 6 linkages in pullulan and α-1, 4 linkages in other polysaccharides, each being maximally active at an alkaline pH, to generate oligosaccharides. We analyzed proteolytic fragments that were produced by exposing pure APase to various proteases, to identify its catalytic domain(s). The intact, pure 210-kDa APase was partially digested with papain for a short time, yielding simultaneously two smaller non-overlapping active fragments, designated amylose-hydrolyzing fragment (AHF114, 114kDa) and pullulan-hydrolyzing fragment (PHF102, 102kDa). The two truncated protein fragments, each containing a single catalytic domain, were purified to homogeneity. The purified AHF114 and PHF102 had similar enzymatic properties to the amylase and pullulanase activities, respectively, of intact APase. The partial amino-terminal sequences of APase and AHF114 were both Glu-Thr-Gly-Asp-Lys-Arg-Ile-Glu-Phe-Ser-Tyr-Glu-Arg-Pro and that of PHF102 was Thr-Val-Pro-Leu-Ala-Leu-Val-Ser-Gly-Glu-Val-Leu-Ser-Asp-Lys-Leu. These results were direct evidence that the α-1, 6 and α-1, 4 hydrolytic activities were associated with two different active sites in this novel enzyme. Our alkaline APase is obviously a "biheaded enzyme".

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A novel enzyme, trehalose synthase, was purified from a cell-free extract of Pimelobacter sp. R48 to an electrophoretically homogeneous state by successive chromatographies on DEAE-Toyopearl 650, Butyl-Toyopearl 650, and Mono Q HR5/5 columns. The molecular weight of the enzyme was estimated to be 62, 000 by SDS-polyacrylamide gel electrophoresis, and the enzyme had a pl of 4.6 by gel isoelectrofocusing. The enzyme catalyzed the lconversion of maltose into trehalose by intramoleculac transglucosylation. The enzyme also converted trehalose into maltose but was inactive on other saccharides. The N-terminal amino acid of the enzyme was serine. The optimum pH and temperature were pH 7.5 and 20°C, respectively. The enzyme was stable in the range of pH 6.0-9.0 and up to 30°C for 60 min. The rate of conversion of maltose into trehalose was independent of the maltose concentration. The maximum yield of trehalose from maltose were 81.8%, 80.9%, and 76.7% at 5, 15, and 25°C, respectively. The activity was inhibited by Cu²⁺, Hg²⁺, Ni²⁺, Zn²⁺, and Tris.

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Cyclodextrin glucanotransferase from an alkalophilic Bacillus species produced neohesperidin monoglucoside and a series of its maltooligoglucosides by transglycosylation with neohesperidin as an acceptor and soluble starch as a donor. As the reaction using β-CD as a donor at an alkaline pH was very effective for solubilizing neohesperidin, the amount of glycosides formed was increased. As a result, its amount with β-CD at pH 10 was about 7 times greater than that with soluble starch at pH 5. Neohesperidin monoglucoside was purified from the reaction mixture by glucoamylase and naringinase treatments, an Amberlite XAD-16 column, a Sephadex LH20 column, and HPLC on an ODS column. The structure of the purified monoglucoside was idenitified as 3^G-α-D-glucopyranosyl neohesperidin by FAB-MS, methylation analysis, and ¹H- and ¹³C-NMR. The solubility of neohesperidin monoglucoside in water was approximately 1500 times higher than that of neohesperidin, and the bitterness of the monoglucoside was about 10 times less than that of neohesperidin. In addition, naringin was also glycosylated by the same method as neohesperidin, and its monoglucoside was identified as ³G-α-D-glucopyranosyl naringin. The solubility of naringin monoglucoside in water was also at least 1000 times higher than that of naringin without altering its bitterness.

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N-Acyl-D-glutamate amidohydrolase (D-AGase) from Pseudomonas sp. 5f-1 was inactivated by diethyl pyrocarbonate (DEP). The chemical modification by DEP showed a difference spectrum at 246nm due to the N-carbethoxyhistidine residue. Removal of the carbethoxy group from inactivated enzyme with hydroxylamine restored enzyme activity. The inactivation by DEP proceeded with pseudo-first-order kinetics, and was protected in the presence of the substrate N-acetyl-D-glutamate (Glu), or the competitive inhibitor sodium α-ketoglutarate (α-KGA). These results suggest the presence of an essential histidine residue at or near of the active site of the enzyme.

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Several compounds were isolated from a Chinese mushroom, Huangmo, the heat-dried fruiting body of Hohenbuehelia serotina. They were idenitified as linoleic acid (I), hexadecanoic acid (II), β-sitosterol (IV), benzoic acid (V), D-mannitol (VI), sucrose (VII), and L-rhamnose (VIII). In addition, six acidic substances were idenitified. (Table I). Also, ethyl linoleate, hexadecanoic acid, and 9, 12-octadecadienoic acid (Z-Z) ethyl esters, IV, V, and VI were idenitified for the first time from this mushroom.

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A novel natural product exhibiting biological activity was isolated from a strain of Botrytis cinerea that had infected raspherry fruit (Rubus ideaus). Liquid fermentation and bioassay-directed fractionation of the organism yielded a compound with molecular formula C₂₂H₃₈O₈ that is trivially named homobotcinolide. It significantly inhibited etiolated wheat coleoptile growth. Green-house-grown bean, corn, and tobacco plants were also affected by exogenous applications of homobotcinolide, severe chlorosis and necrosis being exhibited in corn. The compound is a polyhydroxyl-ated nonalactone esterified with 4-hydroxy-2-decenoic acid.

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The effects of drinking deaerated water on serum biochemical values, and on the concentrations of short-chain fatty acids (SCFAs)derived from bacterial fermentation in the colon were examined in rats. Driniking deaearted water decreased the levels of serum alkaline phosphatase (SAP) and serum urea nitrogen (SUN), and increased the serum potassium (SK) and serum phosphorus (SP) levels. Although the concentration of propionic acid in the cecum was decreased by drinking deaerated water, the concentrations of isobutyric, valeric, and isovaleric acids in the cecum were increased.

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Urea-denatured α-zein was almost completely hydrolyzed into small peptides by digestion with 1/100 (w/w) of thermolysin at 37°C for 3h. The angiotenisin I-converting enzyme (ACE) inhibitory activity (IC₅₀ of the thermolysin digest of total α-zein was 24.5μg/ml, and most of the peptide fractions from Z19 α-zein and total α-zein separated by reverse-phase HPLC had more or less ACE inhibitory activity. From these fractions, thirty-six peptides, including 5 dipeptides, 14 tripeptides, 9 tetrapeptides, 5 pentapeptides, and 3 hexapeptides, were purified and their amino acid sequences were determined.

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The antimicrobial substances in sunflower (Helianthus annuus)were investigated. Two antifungal benzopyran derivatives, 6-acetyl-2, 2-dimethyl-1, 2-benzopyran (1) and 6-acetyl-7-hydroxy-2, 2-di-methyl-1, 2-benzopyran (2) were isolated f om the ethanol extract of sunflower receptacles. The antimicrobial activities of isolated compounds 1 and 2 and their related compounds (precocenes) were evaluated by the paper disk method, using Pyrircularia oryzae as the test fungus.

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We purified an enzyme hydrolyzing 2-sulfo-α-L-fucopyranose pyridylaminiated 2-sulfo-α-L-fucopyranosyl-(1→2)-fucose from the acetone powder of the digestive tract of a sea urchin. The enzyme was purified 307-fold with an overall recovery of 1.63% by ammonium sulfate precipitation, cation exchange chromatography, and gel filtration chromatography. The enzyme is useful for the structural analysis of sulfated fucan.

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Changes in the sensitivity and responsiveness to CCK and cholinergic stimuli were examined in isolated pancreatic acini prepared from 7-day BPJ-diverted rats. The sensitivity to CCK was lower in BPJ-diverted rats than in the control rats, but the sensitivity to the cholinergic agonist, carbachol, was not changed. These results suggest that the contribution of the cholinergic pathway to pancreatic enhancement by feeding dietary protein can be increased after chronic BPJ diversion.

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A uridine-overproducing mutant of Bacillus subtilis was found to require homoserine for growth due to a defect of homoserine dehydrogenase. When the mutant reverted back to prototropy, the production of uridine was decreased by about 40%; the decrease was, however, restored to an appreciable extent by adding aspartic acid, the common precursor for the biosynthesis of homoserine and UMP.

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An endo-1, 4-β-D-mannanase (EC 3.2.1.78) was purified from viscera of a marine mollusc, Littorina brevicula. The purified enzyme, with a molecular weight of 42, 000, was homogeneous by SDS-PAGE. The amino-terminal sequence starting with Gly was analyzed up to the 30th amino acid. The enzyme was stable from pH about 4.0 to about 9.0 and had its maximum activity at pH about 6.5. The purified enzyme produced M2, M3, M4, and M5 from Codium β-1, 4-mannan. The enzyme activity was greatly inhibited by Ag⁺, Hg²⁺, Cu²⁺, and N-bromosuccinimide at 1 mM concentration.

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Stereoisomers and analogues of LL-P880γ (2) were synthesized and tested to elucidate its structure-activity relationship. Their evaluation in the gibberellin-synergistic assay with rice seedlings revealed a clear dependence of potency on the stereochemistry at C1′on the side chain of 2.

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The synthesis of D-2-amino-5-phosphono-3-pentenoic acid (1) is reported. The key intermediate, a (2R, 3S)-3-hydroxyallylglycine derivative (8), was prepared by the reaction of (5S)-3, 6-dimethoxy-5-isopropyl-2, 4-dihydropyrazine (2) and acrolein in the presence of chlorotitanium tris(diethylamide). The transformation of 8 into 1 via allylic alcohol 11 was carried out by following the reported route.

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Optically pure L-p-boronophenylalanine (BPA) was synthesized by a hybrid process involving enantioselective alkylation and subsequent enzymatic hydrolysis. Lithiated (2R)-(-)-2, 5-dihydro-2-isopropyl-3, 6-dimethoxypyrazine was reacted with 4-bromo-methylbenzeneboronate (2) to yield adduct 4 in a 74% e.e. Stepwise treatment of 4 with hydrochloric acid gave L-BPA methyl ester 6, which was hydrolyzed with chymotrypsin to furnish optically pure L-BPA.

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Stevia leaves (Stevia rebaudiana Bertoni; Compositae) accumulate sweet glycosides, the aglycone of which is a diterpene derivative, steviol. The activity of the rate-limiting enzyme of the isoprenoid pathway leading to steviol, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase was found to be extremely (over 150-fold in the specific activity) higher in chloroplasts of stevia leaves than the activity of Spinacia oleracea and about 10-fold higher than that of Solidago altissima L., which also belongs to the Compositae family. There was no difference in apparent K_m values for HMG-CoA and inhibitionis by mevastatin between the activities of thylakoid membranes from S. rebaudiana and S. altissima L. Those activities were almost the same level in microsomal fractions of the three plant leaves.

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Alpha-D-mannosidase was purified from the extract of seeds of Kaya, Torreya nucifera. The purified enzyme had a molecular mass of ∼3.6 x 10⁵ daltons. This enzyme had an optimum pH at 4.5, and was stable at pH between 5.5 and 6.5. This enzyme appeared to be a metal enzyme containing Zn²⁺. The enzyme hydrolyzed p-nitrophenyl-α-D-mannoside, methyl-α-D-mannoside, α-1→3-man-nobiose, and α-1→6-mannobiose, with K_m of 0.785 mM, 0.236 M, 2.505 mM, and 0.268 mM, fespectively. The hydrolysis of various α-linked mannobioses indicated that the enzyme hydrolyzes the α-mannobioses in the order of α-(1→2)> -(1→6)> -(1→3).

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Sphingomonas paucimobilis UT26 uses γ-hexachlorocyclohexane (γ-HCH) as a sole source of carbon and energy. In UT26, γ-HCH is converted to 2, 5-dichlorohydroquinone (2, 5-DCHQ). To clone a gene that is involved in the degradation of 2, 5-DCHQ, a Tn5 mutation was introduced into UT26, and three mutants (UT102, UT103, and UT116) defective in 2, 5-DCHQ degradation were visually selected by spraying γ-HCH solution on the colonies. 2, 5-DCHQ and hydroquinone (HQ) degradation activities of these three mutants and UT26 were analyzed using resting cells. The three mutants had different phenotypes. It was also demonstrated that 2, 5-DCHQ and HQ degradation activities were inducibly expressed in UT26 in the presence of 2, 5-DCHQ.

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6-Phosphogluconate dehydrogenase was screened in for cell-free extracts from seventeen strains of lactic acid bacteria. Three types of the enzyme could be classified according to the coenzyme specificity: NAD-specific, NADP-specific, and non-specific types. The two strains that had a heterofermentative pathway had an NAD-specific 6-phosphogluconate dehydrogenase. This type had higher enzyme activity per proteins in the cell-free extracts, and more specificity to NAD compared to the other types of the enzyme in the lactic acid bacteria studied.

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A transgalactosylation reaction from lactose to moranoline (1-deoxynojirimycin) was accomplished by using β-galactosidase [EC 3.2.1.23] from Bacillus circulans. The enzyme formed 3-O-β-D-galactopyranosyl-moranoline and 4-O-β-D-galactopyranosyl-mor-anolinie as major products, together with 2-O-β-D-galactopranosyl-moranoline and 6-O-β-D-galactopranosyl-moranoline as minor ones.

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Anti-shark type I collagen antibody (PAb-S) was successfully separated from anti-pig type I collagen antiserum. PAb-S recognized a chymotryptic peptide (29 kDa) of α₁ (I) chain prepared from shark skin. This result suggested that the epitope contained in this chymotryptic peptide may be well conserved during the molecular evolution of type I collagen.

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We have isolated 14 cadmium (Cd)-resistant, soil-borne bacteria. Among those, strain CT14, which was identified as an Alcaligenes sp., has a czc (c__-admium, z__-inc, and c__-obalt divalent cation resistant determinant) system. Here we report the nucleotide sequence of 4 genes (czcCBAD) of the system. CzcCBA showed over 98% identity with those of A. eutrophus CH34, however, CzcD, the distal gene product, was 117 amino acids longer than that (199 amino acids) of A. eutrophus CH34, and had considerable similarity to the members of the CDF (cation diffusion facilitator) family proteins all over the region.

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The substrate requirements of coxsackievirus 3C proteinase (3C^pro) were investigated on the C-terminal side of the scissile bond using C-terminal truncated peptides of the substrate peptide Ac-EALFQGPPV. Not only the Gln-Gly bond of Ac-EALFQG-NH₂ but also the C-terminal amide group of Ac-EALFQ-NH₂ was hydrolyzed by 3C^pro, suggesting that the essential residues for cleavage by coxsackievirus 3C^pro would exist within the N-terminal 5 residues.

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Lactobacillus (L.) gasseri JCM1031 has 2 kinds of 6-phospho-β-galactosidase (P-β-gal) in the cytosol. P-β-gal I was already isolated and characterized for our previous paper [Biosci. Biotech. Biochem., 60, 139-141 (1996)]. Another, P-β-gal II, was purified to homogeneity through several liquid chromatographic steps for this paper. The molecular mass of the purified enzyme was estimated to be 58 kDa by SDS-PAGE. The K_m and V_max for ONPGal-6P were 0.76 mM and 86.0 μmol/min/mg, respectively. Reducing reagents and Fe²⁺ ion activated the enzyme, but PCMB, Zn²⁺, and Hg²⁺ ions, and alkylation reagents strongly inihibited it. Twenty-five residues of the N-terminal amino acid sequence of the enzyme were identified. P-β-gal II was different from P-β-gal I in several characteristics.

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Lignin peroxidase of Phanerochaete chrysosporium had a high activity of 3, 3′-dimethoxybenzidine oxidation in 70% aqueous ethylene glycol medium. UV/VIS and ESR spectfoscopic analyses suggested the difference between the oxidation intermediate of 3, 3′-dimethoxybenzidine in water and that in 70% aqueous ethylene glycol medium.

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9-Aminio-2-hydroxyacridine, a rat liver S9 metabolite of 9-aminoacridine (9-AA), was synthesized, and found to have lower frameshift mutagenicity and stronger DNA binding affinity than 9-AA.

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The two genes encoding maltooligosyl trehalose synthase and maltooligosyl trehalose trehalohydrolase, which are related to biosynthesis of α, α-trehalose, were cloned from Rhizobium sp. M-11. Sequence analysis showed that the synthase gene composed of 2316 bp was connected with the hydrolase gene of 1788 bp by an overlap of one nucleotide. The deduced amino acid sequences of both enzymes have several regions common to amylolytic enzymes belonging to an "α-amylase family".

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Transient expression of the chloramphenicol acetyltransferase gene (cat) under the control of Simian virus 40 early enhancer/promoter complex (pAOcat) was stimulated initially by differentiation of F9 cells into primitive endoderm, but it was repressed by further differentiation into visceral endoderm. Deletion of the 72-base pairs of enhancer reduced cat expression, but the dependence on differentiation was still observed. Gel retardation Assays using enhancer or promoter sequenices revealed nuclear factors expressed in undifferentiated (stem) and visceral endoderm F9 cells. Cotransfection of pAOcat with an excess of promoter or enhancer sequence stimulated cat expression. Thus, repressor-like factors were suggested to be responsible for the differentiation-dependent control.

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The structure of cochlioquinol (1), a new compound isolated from a culture of the plant pathogenic fungus, Bipolaris cynodontis, was determined, based on single-crystal X-ray diffraction studies. Compound 1 was found to be related to cochlioquinone derivatives that had previously been isolated as phytotoxins from a culture of B. bicolor, in which the addition of acetone occurred at one of the quinonie carbonyl groups of cochlioquinone. Compound 1 inhibited the root growth of Italian ryegrass, one of the host plants of B. cynodontis, by 50% at a concentration of 100ppm.

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Streptomyces sp. strain No. 6288 produces S-Hemolysin, which is a unique phospholipase C with a high substrate specificity for sphingomyelin. Moreover, the strain also produced two kinds of phospholipase inhibitors, designated as SHI and S-PLI, in different phases of cultivation. The purified SHI was shown to be a basic substance containing an amino group and glycoside moiety, and it was a more effective inhibitor of S-Hemolysin and sphingomyelinase from Streptomyces sp. with a higher substrate specifcity for sphingomyelin than α-toxin from Clostridium perfringens.

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A selective and sensitive method for the determination of glutathione and such related aminothiols as Cys, cysteinylglycine and γ-glutamylcysteine in mouse tissues by gas chromatography was developed. After fractionating a tissue sample into the free and protein-bound fractions, glutathione and aminothiols in each fraction were converted inito their N, S-isopropoxycarbonyl methyl ester derivatives and then measured by gas chromatography with flame photometric detection, using a DB-1 capillary column. This method enabled the redox status of glutathione and related aminothiols in mouse tissues to be investigated.

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Three new glucosidic constituents (1-3) were isolated from the methanolic extract of Perilla frutescens. Two of these compounds (1 and 2) were jasmonoid glucosides and were determined to be 5′-β-D-glucopyranosyloxyjasmonic acid and 3-β-D-glucopyranosyl-3-epi-2-isocucurbic acid, respectively. The other one (3) Was a new glucoside of phenylvaleric acid that was elucidated to be 3-β-D-glucopyranosyloxy-5-phenylvaleric acid. Its aglycone was also found for the first time in nature.

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Cell-free extracts of petioles of Arctium lappa catalyzed enantioselective formation of (+)-secoisolariciresinol [about 20% enantiomer excess (e.e.)] from achiral coniferyl alcohol in the presence of NADPH and H₂O₂. This is the first report of an enzymatic reaction to afford (+)-secoisolariciresinol enantioselectively.

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