(Vol.61 No.7 1997)
-Review-
Preparation of Oligosaccharide Units Library and Its Utilization
Takeomi Murata and Taichi Usui 1059
Random Amplified Polymorphic DNA (RAPD) Analyses for Discriminating
Genotypes of Microcystis Cyanobacteria
Hirofumi Nishihara, Hiroyasu Miwa, Masayuki Watanabe, Minoru Nagashima,Osami
Yagi, and Yoshichika Takamura 1067
Induction of -Glucosidase in Botrytis cinerea by Cell
Wall Fractions of the Host Plant
Izumi Sasaki and Hideo Nagayama 1073
Structural Clarification of Caulerpa Cell Wall -1,3-Xylan
by NMR Spectroscopy
Tohru Yamagaki, Masaakira Maeda, Kenji Kanazawa, Yasuko Ishizuka, and
Hiroshi Nakanishi 1077
Purification and Some Properties of GTP Cyclohydrolase
I from Spinach Leaves
Yasuko Sohta, Tomoko Ohta, and Masahiro Masada 1081
Purification and Calcium Dependence of Transglutaminases
from Sheep Hair Follicles
Yoshiyuki Kumazawa, Tomoko Ohtsuka, Daiki Ninomiya, and Katsuya Seguro
1086
A Catalytic Amino Acid and Primary Structure of Active
Site in Aspergillus niger -Glucosidase
Atsuo Kimura, Masuhiro Takata, Yukiharu Fukushi, Haruhide Mori, Hirokazu
Matsui,and Seiya Chiba 1091
Inhibitory Effects of Persimmon (Diospyros kaki ) Extract
and Related Polyphenol Compounds on Growth of Human Lymphoid Leukemia Cells
Yumiko Achiwa, Hiroshige Hibasami, Hirotaka Katsuzaki, Kunio Imai and
Takashi Komiya 1099
Purification and Properties of an Aminopeptidase from a
Protamine-degrading Marine Bacterium
Hitoshi Obata, Atsushi Sugiyama, Hidehisa Kawahara, and Tsuyoshi Muramatsu
1102
Relationship between the Glutamate Production and the
Activity of 2-Oxoglutarate Dehydrogenase in Brevibacterium lactofermentum
Yoshio Kawahara, Keiko Takahashi-Fuke, Eiko Shimizu, Tsuyoshi Nakamatsu,
and Shigeru Nakamori 1109
Purification and Properties of Two Deacetylases Produced
by Vibrio alginolyticus H-8
Kazuo Ohishi, Masaaki Yamagishi, Toshiya Ohta, Masayoshi Motosugi,
Hitoshi Izumida,Hiroshi Sano, Kyoko Adachi, and Tan Miwa 1113
Galactosyl Transfer onto p-Nitrophenyl -d-Glucoside
Using -d-Galactosidase from Bacillus circulans
Takeomi Murata, Satoru Akimoto, Miki Horimoto, and Taichi Usui 1118
The Effects of Substituents Introduced into 9-Aminoacridine
on Frameshift Mutagenicity and DNA Binding Affinity
Hideyuki Tomosaka, Saburo Omata, Eietsu Hasegawa, and Kentaro Anzai
1121
Further Studies on Thermal Denaturation of Pyruvate Dehydrogenase
Complex from Bacillus stearothermophilus
Yasuaki Hiromasa, Yoichi Aso, Shoji Yamashita, Yoshikatsu Aikawa, and
Masatsune Ishiguro 1126
Inactivation of Food Microorganisms by High-pressure Carbon
Dioxide Treatment with or without Explosive Decompression
Atsushi Enomoto, Kozo Nakamura, Kiyotaka Nagai, Takeki Hashimoto, and
Masaru Hakoda 1133
Isolation and Activity of N-p-Coumaroyltyramine, an -Glucosidase
Inhibitor in Welsh Onion (Allium fistulosum)
Tetsuo Nishioka, Jun Watanabe, Jun Kawabata, and Ryoya Niki 1138
Isomeric Ratio and Level of Xanthoxin in Tomato Plants
Measured by a New Analytical Method
Hirotaka Yamamoto and Takayuki Oritani 1142
Synthesis of Glycosyl-trehaloses by Cyclomaltodextrin
Glucanotransferase through the Transglycosylation Reaction
Masashi Kurimoto, Akihiko Tabuchi, Takahiko Mandai, Takashi Shibuya,
Hiroto Chaen,Shigeharu Fukuda, Toshiyuki Sugimoto, and Yoshio Tsujisaka
1146
Effects of Medium-chain Fatty Acids and Their Acylglycerols
on the Transport of Penicillin V across Caco 2 Cell Monolayers
Motohiro Shima, Kaori Yohdoh, Masayo Yamaguchi, Yukitaka Kimura, Shuji
Adachi, Ryuichi Matsuno 1150
Enhancing Effect of Interleukin-4 on the Secretion of
Interferon- by (/)s1-Casein-specific CD8+ T Cells
Ken-ichi Nishijima, Masako Kohyama, Tatsuhiro Hisatsune, Hiroko Kato,
Masahiro Kakehi,and Shuichi Kaminogawa 1156
Further Studies on the Preparation of Low Sodium Chloride-containing
Soy Sauce by Using Ornithyltaurine Hydrochloride and Its Related Compounds
Rie Kuramitsu, Daisuke Segawa, Kozo Nakamura, Shunsuke Muramatsu, and
Hideo Okai 1163
Purification and Characterization of Glutamate Decarboxylase
from Lactobacillus brevis IFO 12005
Yoshie Ueno, Kiyoshi Hayakawa, Saori Takahashi, and Kohei Oda 1168
Isolation and Characterization of kar2-404 Mutation in
Saccharomyces cerevisiae
Akiko Kawamura-Watabe and Masao Tokunaga 1172
Separation and Determination of Yellow and Red Safflower
Pigments in Food by@Capillary Electrophoresis
Toshiro Watanabe, Naoki Hasegawa, Akira Yamamoto, Shiro Nagai, and
Shigeru Terabe 1179
-note-
Purification and Characterization of a Glycine-rich Polypeptide Tightly
Bound to Cell Walls from Soybean Aleurone Layers
Mitsuru Fukuda, Hiroko Noda, and Isao Toyosawa 1184
Characteristics of Escherichia coli HB101 and Pseudomonas
putida PpY101 Harboring a Recombinant Plasmid with Tandem Insertion of
the Mercury Resistance Operon
Teruyo Kurabayashi, Kazuhiro Iwasaki, Hiroo Uchiyama, Kunihiko Nakamura,Hideo
Tanaka, and Osami Yagi 1187
Distribution of Cu-resistant and Non-resistant Bacteria
in Several Cu-contaminated Soils in Japan: Usefulness of the Tolerance
Level as an Index of Cu Contamination in Soil
Takashi Kunito, Keishi Senoo, Kazunari Nagaoka, Hiroshi Oyaizu, and
Satoshi Matsumoto 1190
Effect of Chaotropic Salt on the Secondary Structure of
Pigskin Gelatin
Yong Sik Cho and Kyung Bin Song 1194
Increase of Catalytic Activity of -Chymotrypsin by Metal
Salts for Transesterification of an Amino Acid Ester in Ethanol
Toshiya Sasaki and Hideo Kise 1196
Fluorescence-labeled Abscisic Acid Possessing Abscisic
Acid-like Activity in Barley Aleurone Protoplasts
Tadao Asami, Ling Tao, Shin Yamamoto, Masumi Robertson, Yong-Ki Min,Noboru
Murofushi, and Shigeo Yoshida 1198
Effects of Sex Hormones on the Metabolism of Tryptophan
to Niacin and to Serotonin in Male Rats
Katsumi Shibata and Satoko Toda 1200
Chemical Characterization of a Sulfated Polysaccharide-Peptidoglycan
Complex from an Arthrobacter sp.
Ken-ichi Yamazaki, Makoto Suzuki, Kazuyoshi Inukai, Hideo Hakusui 1203
Accelerating Effect of Chitosan Intake on Urinary Calcium
Excretion by Rats
Masahiro Wada, Yoshikazu Nishimura, Yoshito Watanabe, Toshichika Takita,
Satoshi Innami 1206
Effect of Phosphoryl Oligosaccharides on Iron Solubility
under Neutral Conditions
Hiroshi Kamasaka, Kenji To-o, Kaname Kusaka, Takashi Kuriki, Takashi
Kometani, Shigetaka Okada 1209
Construction of Escherichia coli-Bifidobacterium longum
Shuttle Vector Transforming B. longum 105-A and 108-A
Hajime Matsumura, Akio Takeuchi, and Yasunobu Kano 1211
Synthesis and Insecticidal Activity of New 2- and 6-Substituted
4-Acetoxyquinolines
Nobuto Minowa, Kei-ichi Imamura, and Seiji Shibahara 1213
A New Enzymatic Method for L-Phenylalanine Synthesis Using
Aminoacylase andPhenylalanine Dehydrogenase
Hong-Yon Cho and Kenji Soda 1216
Solubilizing Coelenterazine in Water with Hydroxypropyl--cyclodextrin
Katsunori Teranishi and Osamu Shimomura 1219
-Rapid Paper-
Thiamine Increases Expression of Yeast Gene
Kimihisa Ichikawa, Yoichiro Shiba, Mitsuo Yamazaki, and Nobufusa Serizawa
1221
An Anti-hydrotactic Response and Solid Surface Recognition
of Germ Tubes of the Rice Blast Fungus, Magnaporthe grisea
Jin-zhong Xiao, Tadakazu Watanabe, Shigeko Sekido, Woo-Bong Choi, Takashi
Kamakura, and Isamu Yamaguchi 1225
Conformational Behavior of Chitosan in the Acetate Salt:
An X-Ray Study
Atsushi Yamamoto, Junpei Kawada, Toshifumi Yui, and Kozo Ogawa 1230
-1-
Review
uPreparation of Oligosaccharide Units Library and Its Utilizationv
Takeomi Murata and Taichi Usui
Department of Applied Biological Chemistry, Faculty of Agriculture, Shizuoka University, 836 Ohya, Shizuoka 422, Japan
@There is high current interest in developing synthetic routes to oligosaccharides
involved in glycoconjugates. Significant attention has been focused on
the application of glycosidase-catalyzed transglycosylation for practical
synthesis of oligosaccharides. The enzymatic synthesis has become more
practical by the use of several glycosidases available in sufficient quantities.
This review describes convenient syntheses of di- and trisaccharide units,
which are related to molecular recognition, by using regioselective transgalactosylation,
trans-N-acetylglucosaminylation, transfucosylation, and transmannosylation.
The regioselectivity could be controlled to some extent by using the following
techniques: (1) varying enzymes, (2) organic co-solvent system, (3) the
configuration of the existing glycosidic linkage of the acceptor and (4)
inclusion complex of acceptor glycoside with cyclodextrin. Furthermore,
glycopolymers carrying a series of disaccharides containing -D-galactosyl
residues were synthesized and used as a model in oligosaccharide-lectin
interaction analysis. These water-soluble glycopolymers were shown to be
useful as probes of carbohydrate recognition.
Key words: oligosaccharide units library; glycosidase-catalyzed transglycosylation;
glycoconjugate; glycopolymer
-2-
Random Amplified Polymorphic DNA (RAPD) Analyses for Discriminating
Genotypes of Microcystis Cyanobacteria
Hirofumi Nishihara, Hiroyasu Miwa, Masayuki Watanabe,* Minoru Nagashima,** Osami Yagi,*** and Yoshichika Takamura
Department of Applied Biological Resource Sciences, Ibaraki University,
3-21-1 Chu-ou, Ami-machi, Ibaraki 300-03, Japan
* Department of Botany, National Science Museum, 4-1-1 Amakubo, Tsukuba-shi,
Ibaraki 305, Japan
** Tsukuba Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., 2 Miyukigaoka,
Tsukuba-shi, Ibaraki 305, Japan
*** Water and Soil Environment Division, The National Institute for Environmental
Studies, 16-2 Onogawa, Tsukuba-shi, Ibaraki 305, Japan
Received September 2, 1996
@Random amplified polymorphic DNA (RAPD) analysis was used to discriminate
genotypes in five species of Microcystis cyanobacteria. Strains of each
group with the identical allozyme genotype (T. Kato et al., Algol. Stud.,
1991, 129-140; M. Watanabe, in eeToxic Microcystis,'' ed. by M. F. Watanabe
et al., CRC Press, Tokyo, 1966, pp. 13-34) gave similar RAPD patterns characterizing
the respective group. On the other hand, no similarities in RAPD patterns
were observed among strains of which allozyme genotypes were different.
A good accordance between the RAPD analysis and allozyme divergence indicated
a high reliability of both methods for discrimination of the affiliated
groups of Microcystis. Several amplified DNA fragments, which were expected
to be markers for a particular taxon with identical allozyme genotype,
were also observed on the RAPD patterns. Genetic homogeneities of M. novacekii,
M. viridis, and M. wesenbergii were shown by RAPD analysis as well as the
allozyme genotype. However, significant variations were observed in M.
aeruginosa and M. ichthyoblabe in the levels of DNA and proteins (allozymes).
Key words: RAPD analysis; DNA polymorphism; Microcystis; cyanobacterium;
allozyme genotype
-3-
Induction of -Glucosidase in Botrytis cinerea by Cell
Wall Fractions of the Host Plant
Izumi Sasaki and Hideo Nagayama*
Department of Chemistry and Bioengineering, Oyama National College of
Technology, Oyama 323, Japan
* Department of Science and Technology, Ishinomaki Senshu University, Ishinomaki
986, Japan
Received September 18, 1996
@The pathogenicity of Botrytis cinerea has been found to correlate positively
with the -glucosidase activity. In this report, the relationship between
the induction of -glucosidase and the components of host plant tissues
was studied by the use of tissue fractions and cellulose-related compounds.
@The most active enzyme induced by the crude fiber fraction and Avicel
was -glucosidase, among the cell wall degrading enzymes tested. The -glucosidase
was very inducible in the strains with strong pathogenicity, and intensively
degraded the fiber fraction made from apple fruit tissues. The same degradation
of the cell wall fraction was demonstrated with the purified enzyme.
Key words: inducible enzyme; -glucosidase; Botrytis cinerea; phytopathology
-4-
Structural Clarification of Caulerpa Cell Wall -1,3-Xylan
by NMR Spectroscopy
Tohru Yamagaki, Masaakira Maeda, Kenji Kanazawa,* Yasuko Ishizuka,* and Hiroshi Nakanishi *
Department of Biochemistry, Saitama University, Urawa, Saitama 338,
Japan
* National Institute of Bioscience and Human-Technology, AIST, 1-1 Higashi,
Tsukuba, Ibaraki 305, Japan
Received September 19, 1996
@1H- and 13C-NMR spectroscopic analyses of cell wall microfibril -1,3-xylan
from Caulerpa brachypus were performed in detail. The total assignment
of all 1H- and 13C-NMR signals in the -1,3-xylan was achieved by evaluating
the 2D C-H COSY, DQF-COSY, and NOESY spectra. The results, including information
on through-space interaction between the xylopyranose residues, were confirmed.
The determination of the glycosidic linkages by NMR spectroscopic analyses
agrees well with the results from a chemical analysis.
Key words: Caulerpa brachypus ; -1,3-xylan; NMR spectroscopy; NOE
-5-
Purification and Some Properties of GTP Cyclohydrolase
I from Spinach Leaves
Yasuko Sohta, Tomoko Ohta, and Masahiro Masada
Department of Bioresources Chemistry, Faculty of Horticulture, Chiba University, Matsudo 648, Chiba 271, Japan
Received October 2, 1996
@GTP cyclohydrolase I (EC 3.5.4.16) has been purified for the first time
from a higher plant, spinach leaves. The purified preparation appeared
to be homogeneous on polyacrylamide gel electrophoresis. The molecular
weight of this enzyme was estimated at 135,000 by gel filtration and the
subunit molecular weight was estimated at 35,000 by SDS-PAGE. The latter
method also suggested that this enzyme was composed of four identical subunits.
The enzyme was stable to heat treatment at 50C for 10 min, and the activity
was maintained for at least six months when stored at -30C. The enzyme
had an optimum pH of around 8.0 in Tris buffer. The Hill coefficient of
the enzyme was calculated to be 2.2. The pI of the enzyme was measured
as 5.1 by chromatofocusing.
Key words: GTP cyclohydrolase I; spinach; purification; characterization
-6-
Purification and Calcium Dependence of Transglutaminases
from Sheep Hair Follicles
Yoshiyuki Kumazawa, Tomoko Ohtsuka, Daiki Ninomiya, and Katsuya Seguro
Ajinomoto Co., Inc., Food Development & Research Laboratories, 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki-shi, Kanagawa 210, Japan
Received October 4, 1996
@To study the calcium sensitivity of sheep hair follicle transglutaminase,
which was reportedly calcium-independent [H. W. Harding and G. E. Rogers,
Biochemistry, 11, 2858-2863 (1972)], the enzyme was purified from a homogenate
of merino sheep hair follicles and its calcium dependence was examined.
As a result of purification, two types of transglutaminases (DEAE-unabsorbed
and absorbed transglutaminase, DU-TG and DA-TG, respectively) were obtained.
The molecular mass of DU-TG was 77 and 82 kDa by SDS-PAGE and gel filtration,
respectively, while that of DA-TG was 40 and 80 kDa. Each enzyme was obviously
calcium dependent and contained (a) cysteine residue(s) in the active site,
like other known mammalian transglutaminases. Maximum activation of DU-TG
and DA-TG was observed at 1 and 0.1 mM CaCl2, respectively.
Key words: purification; sheep hair follicles; transglutaminase; calcium-dependency
-7-
A Catalytic Amino Acid and Primary Structure of Active
Site in Aspergillus niger -Glucosidase
Atsuo Kimura, Masuhiro Takata, Yukiharu Fukushi, Haruhide Mori, Hirokazu Matsui,* and Seiya Chiba
Department of Applied Bioscience, Faculty of Agriculture, Hokkaido University,
Sapporo 060, Japan
* Department of Bioscience and Chemistry, Faculty of Agriculture, Hokkaido
University, Sapporo 060, Japan
Received November 15, 1996
@The catalytic amino acid residue of Aspergillus niger -glucosidase
(ANGase) was identified by modification with conduritol B epoxide (CBE),
a mechanism-based irreversible inactivator. The inactivation by CBE followed
pseudo-first order kinetics. The interaction of CBE and ANGase conformed
to a model with a reversible enzyme-inhibitor complex formed before covalent
inactivation. A competitive inhibitor, Tris, decreased the inactivation
rate. The incorporation of one mole of CBE per mole of ANGase was completely
abolished the enzyme activity. A dissociated carboxyl group (-COO-) in
the active site was suggested to attack the C-1 of CBE. ANGase was composed
of two subunits (P1 and P2), of which P2 was modified by CBE. The labelled
residue was included in a peptide (LY3) that was obtained from Lys-C protease
digestion of CBE-bound P2. The sequence analysis of CBE-labelled LY3 showed
that an Asp was the modified residue, that is, one of the catalytic amino
acid residues of ANGase. The primary structure of LY3 was determined by
analyzing the sequence of peptide fragments prepared by several proteases.
Key words: catalytic amino acid; -glucosidase; Aspergillus niger; primary
structure; affinity labelling
-8-
Inhibitory Effects of Persimmon (Diospyros kaki ) Extract
and Related Polyphenol Compounds on Growth of Human Lymphoid Leukemia Cells
Yumiko Achiwa,* Hiroshige Hibasami,** Hirotaka Katsuzaki,* Kunio Imai,* and Takashi Komiya*,
* Faculty of Bioresources and ** College of Medical Science, Mie University, Tsu-shi, Mie 514, Japan
Received November 19, 1996
@We have investigated the effects of persimmon (Diospyros kaki ) extract
(PS) and related polyphenol compounds such as catechin (C), epicatechin
(EC), epicatechingallate (ECG), epigallocatechin (EGC), and epigallocatechingallate
(EGCG) on the growth of human lymphoid leukemia Molt 4B cells. We found
that PS, ECG, EGC, and EGCG strongly inhibited the growth of the cells
in a dose-dependent manner, while C and EC inhibited the growth of the
cells only moderately. Ornithine decarboxylase (ODC), a rate-limiting enzyme
of polyamine biosynthesis, was inhibited by 10-20% by these polyphenol
compounds. The morphology of the Molt 4B cells indicated severe damage
3 days after treatment with PS, ECG, EGC, and EGCG. Irregular shape of
the cells and DNA fragmentation were observed in PS, ECG, EGC, or EGCG-treated
cells. These results suggest that PS, ECG, EGC, and EGCG induce apoptosis
(programmed cell death) of Molt 4B cells.
Key words: persimmon extract; polyphenol compounds; apoptosis; leukemia
-9-
Purification and Properties of an Aminopeptidase from a
Protamine-degrading Marine Bacterium
Hitoshi Obata,*,**, Atsushi Sugiyama,* Hidehisa Kawahara,*,** and Tsuyoshi Muramatsu ***
* Department of Biotechnology, Faculty of Engineering, Kansai University,
Yamatecho 3-3-35, Suita-shi, Osaka 564, Japan
** Kansai University High Technology Research Center, Yamatecho 3-3-35,
Suita-shi, Osaka 564, Japan
*** Faculty of Fisheries, Nagasaki University, Nagasaki-shi, Nagasaki 852,
Japan
Received November 22, 1996
@A protamine-degrading marine bacterium was isolated from marine soil
and identified as Aeromonas salmonicida subsp. based on its taxonomical
characteristics. An alanine-specific aminopeptidase, called aminopeptidase
K, from an extract of the strain was purified and characterized. The aminopeptidase
K was purified about 80-fold by fractionation with ammonium sulfate and
column chromatography on QA-52 cellulose, Phenyl Superose and Superose
12. The purified enzyme is composed of 6 subunits of 86 kDa with a molecular
mass of 520 kDa according to gel filtration and SDS-PAGE. The N-terminal
sequence of the enzyme was HEGly-Gln-Gln-Pro-Gln-Ile-Lys-Try-Tyr-His-Asp-Tyr-Asp-Ala-Pro-Asp-Tyr-Tyr-Ile-Thr-.
It is inhibited by monoiodoacetate, N-ethylmaleimide, and puromycin. The
Michaelis constant (Km) and the maximal rate of hydrolysis (V(/)max) were,
respectively, 0.28 mM and 49.4 mol/min/mg for the L-Ala--naphthylamide
substrate. The optimum pH and optimum temperature were 6.5 and 45C, respectively.
The purified enzyme was highly specific to L-Ala--naphthylamide.
Key words: marine bacterium; aminopeptidase; Aeromonas salmonicida; alanine
aminopeptidase; l-Ala--naphthylamide
-10-
Relationship between the Glutamate Production and the
Activity of 2-Oxoglutarate Dehydrogenase in Brevibacterium lactofermentum
Yoshio Kawahara, Keiko Takahashi-Fuke, Eiko Shimizu, Tsuyoshi Nakamatsu, and Shigeru Nakamori
Technology and Engineering Laboratories of Ajinomoto Co., Inc., 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki 210, Japan
Received December 2, 1996
@Enzyme activities of 2-oxoglutarate dehydrogenase complex and glutamate
dehydrogenase of wild type Brevibacterium lactofermentum, one of the typical
glutamate-producing coryneform bacteria, were investigated by using cells
cultured under glutamate-productive and glutamate-non-productive conditions.
@Significant reduction of the former enzyme activity was observed in the
cells under the several glutamate-productive conditions, namely, in the
cells cultured in media containing a) limited concentrations of biotin,
b) sub-lethal amounts of penicillin, and c) sub-optimal amounts of a surface-active
agent, as compared with those under the non-productive conditions. The
activity of the latter enzyme was essentially unchanged in every condition.
@The relationship between glutamate production and the enzyme activities
as well as permeability of glutamate through cell membrane was discussed
from the results obtained.
Key words: glutamate production; Brevibacterium lactofermentum; 2-oxoglutarate
dehydrogenase
-11-
Purification and Properties of Two Deacetylases Produced
by Vibrio alginolyticus H-8
Kazuo Ohishi, Masaaki Yamagishi, Toshiya Ohta, Masayoshi Motosugi, Hitoshi Izumida,* Hiroshi Sano,* Kyoko Adachi,* and Tan Miwa**
Numazu Industrial Research Institute of Shizuoka Prefecture, 3981-1
Ohoka, Numazu, Shizuoka 410, Japan
* Marine Biotechnology Institute Co., Ltd., 1900 Sodeshi-cho, Shimizu,
Shizuoka 424, Japan
** KEI Chemical Industry Co., Ltd., 328 Shioshinden-hamano, Fukude-cho,
Iwata-gun, Shizuoka 437-12, Japan
Received December 2, 1996
@The Chitinase-producing bacterium Vibrio alginolyticus H-8 isolated from
mud of Hamana Lake also produced two deacetylases for (GlcNAc)2 extracellularly.
Deacetylases DA1 and DA2 were purified from crude enzyme by column chromatography
on Q-Sepharose FF, Phenyl Sepharose HP, Gigapite, and Superdex 200 HR.
The final preparation was homogeneous in SDS-PAGE. The molecular weights
were 48,000 and 46,000 for deacetylases DA1 and DA2, respectively. The
pIs, optimum pHs, and optimum temperatures for deacetylases DA1 and DA2
were as follows; DA1, pI 3.3, optimum pH 8.5-9.0, optimum temperature 45C,
DA2, pI 3.5, optimum pH 8.0-8.5, optimum temperature 40C. Both deacetylases
were stable at pHs between 7.0 and 11.0 and at temperatures below 40C.
The activities of both enzymes were inhibited by Ag+ and Hg2+. 1H-NMR of
the reaction product by deacetylase DA1 for (GlcNAc)2 showed that the purified
deacetylase selectively hydrolyzed the 2-acetamide group at the reducing
end of (GlcNAc)2.
Key words: deacetylase; N-monoacetylchitobiose; N,N '-diacetylchitobiose;
chitin; Vibrio alginolyticus
-12-
Galactosyl Transfer onto p-Nitrophenyl -D-Glucoside
Using -D-Galactosidase from Bacillus circulans
Takeomi Murata, Satoru Akimoto, Miki Horimoto, and Taichi Usui *
Faculty of Agriculture, Department of Applied Biological Chemistry, Shizuoka University, 836 Ohya, Shizuoka 422, Japan
Received December 9, 1996
@-D-Gal-(14)--D-Glc-OC6H4NO2-p and its isomers (-D-Gal-(13)--D-Glc-OC6H4NO2-p
and -D-Gal-(16)--D-Glc-OC6H4NO2-p) were synthesized from lactose
and -D-Glc-OC6H4NO2-p, using transglycosylation by the -D-galactosidase
from Bacillus circulans. This reaction was efficient enough for us to do
a one-pot preparation of galactosyl-glucoside from lactose. The order of
the production of the transfer products was (14) (13)>(16)
in the initial stage of the reaction, and the same relationship was observed
for the hydrolytic rate toward the three galactosyl-glucosides. The production
of (14)- and (13)-linkages greatly decreased during the subsequent
reaction and much more of the (16)- than of the (14)- and (13)-transfer
products was found in the later stage of the reaction.
Key words: -d-galactosidase; galactosyl-glucoside; enzymic synthesis;
transglycosylation; regioselectivity
-13-
The Effects of Substituents Introduced into 9-Aminoacridine
on Frameshift Mutagenicity and DNA Binding Affinity
Hideyuki Tomosaka, Saburo Omata, Eietsu Hasegawa,* and Kentaro Anzai**
Department of Biosystem Science, Graduate School of Science and Technology,
and * Department of Chemistry, Faculty of Science, Niigata University,
Igarashi, Niigata, Niigata 950-21, Japan
** Department of Chemistry, Faculty of Science, Science University of Tokyo,
Kagurazaka, Shinjuku-ku, Tokyo 162, Japan
Received December 16, 1996
@Some derivatives of 9-aminoacridine (1) were synthesized, and their frameshift
mutagenicity and DNA binding affinity were studied. The introduction of
a methyl group into the acridine ring of 1 reduced the mutagenic activity
and the intercalative DNA binding affinity, while the introduction of chlorine
increased them. Halogenated derivatives of 1 showed higher toxicity against
Salmonella typhimurium TA1537.
Key words: Ames test; mutagenicity; DNA intercalation; Scatchard analysis;
acridine
-14-
Further Studies on Thermal Denaturation of Pyruvate Dehydrogenase
Complex from Bacillus stearothermophilus
Yasuaki Hiromasa, Yoichi Aso, Shoji Yamashita,* Yoshikatsu Aikawa, and Masatsune Ishiguro
Laboratory of Protein Chemistry and Engineering, Graduate School of Genetic Resources Technology, and * Institute of Biophysics, Faculty of Agriculture, Kyushu University, Fukuoka 812-81, Japan
Received December 18, 1996
@Thermally induced changes in pyruvate dehydrogenase complex (PDC) from
B. stearothermophilus were examined mainly at temperatures from 60 to
70C. Accompanied by inactivation of pyruvate decarboxylase, light scattering
decreased, and ANS fluorescence increased. These changes including the
inactivation were approximately first-order reactions, and the values of
rate constants were greatly dependent on termperature. Chromatographic
studies showed that any polypeptides were in associated forms and that
final products were aggregates 30S) and an assembly (48S) smaller than
PDC. The aggregates and assembly were rich in decarboxylase and lipoate
acetyltransferase, respectively.
@It was suggested that, during the thermal denaturation, a decarboxylase
was dissociated from PDC and immediately involved in aggregates.
Key words: Bacillus stearothermophilus; denaturation; multienzyme complex;
pyruvate dehydrogenase
-15-
Inactivation of Food Microorganisms by High-pressure Carbon
Dioxide Treatment with or without Explosive Decompression
Atsushi Enomoto, Kozo Nakamura,* Kiyotaka Nagai, Takeki Hashimoto, and Masaru Hakoda
Department of Biological and Chemical Engineering, Faculty of Engineering,
Gunma University, 1-5-1 Tenjin, Kiryu, Gunma 376, Japan
* Department of Applied Biological Chemistry, Graduate School of Agriculture
and Agricultural Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku,
Tokyo 113, Japan
Received December 19, 1996
@In order to elucidate the sterilization mechanism underlying the explosive
decompression system, baker's yeast was pressurized with CO2, N2O, N2,
or Ar gas at 40 atm and 40C for 4 h, and then explosively discharged.
The survival ratio was markedly decreased only by the treatments with CO2
and N2O, which are relatively soluble gases in water, suggesting that the
microorganisms' death may be highly correlated with gas absorption by the
cells. Lower decompression rates to atmospheric pressure, however, led
to neither any lower reduction of remaining cells nor any smaller release
of total cellular proteins. Furthermore, operating with a longer treatment
time and smaller number of repetitions was usually more lethal than with
a shorter time and more frequent repetition. From these results, most of
the yeast cells appear to have been sterilized during the pressurization
process. The spore cells of B. megaterium are considered to have been killed
in a somewhat different manner, because of their distinct sensitivity to
the applied gases.
Key words: sterilization; high-pressure carbon dioxide; food microorganism;
survival ratio; decompression rate
-16-
Isolation and Activity of N-p-Coumaroyltyramine, an -Glucosidase
Inhibitor in Welsh Onion (Allium fistulosum)
Tetsuo Nishioka, Jun Watanabe, Jun Kawabata, and Ryoya Niki
Department of Bioscience and Chemistry, Faculty of Agriculture, Hokkaido University, Kita-ku, Sapporo 060, Japan
Received December 20, 1996
@A phenolic amide, N-p-coumaroyltyramine (1), was isolated as an -glucosidase
inhibitor from methanol extracts of Welsh onion (Allium fistulosum). The
inhibitory activity of 1 against a yeast enzyme was as high as Ki 8.4~10-7
M. From a structure-activity relationship study of 1 and its related compounds,
the occurrence of -glucosidase inhibitory activity required a p-coumaramide
structure, with an amide hydrogen and alkyl or aralkyl substituent on the
amide part.
Key words: -glucosidase inhibitor; N-p-coumaroyltyramine; Allium fistulosum
-17-
Isomeric Ratio and Level of Xanthoxin in Tomato Plants
Measured by a New Analytical Method
Hirotaka Yamamoto and Takayuki Oritani
Department of Applied Biological Chemistry, Faculty of Agriculture, Tohoku University, 1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai 981, Japan
Received December 24, 1996
@The isomeric ratio and level of natural xanthoxin (XAN) in tomato plants
(Lycopersicon esculentum) were examined by a more reliable analytical method
than has been reported before. Efforts were made to avoid artificial isomerization
between c-XAN and t-XAN throughout the isolation, derivatization and GC-MS
procedures. Natural XAN was separated from contaminating chlorophylls before
rev. HPLC purification, derivatized to abscisic acid methyl ester (MeABA)
in four chemical steps, and quantified with the deuterium-labeled internal
standards on clear and reproducible full GC-EI-MS. It was revealed that
the isomeric composition of natural XAN was exclusively shifted to c-XAN.
The level of c-XAN was higher and more significantly induced by water stress
in older plants. The significant role of c-XAN as an ABA biosynthetic precursor
is suggested.
Key words: abscisic acid; biosynthesis; xanthoxin
-18-
Synthesis of Glycosyl-trehaloses by Cyclomaltodextrin
Glucanotransferase through the Transglycosylation Reaction
Masashi Kurimoto, Akihiko Tabuchi, Takahiko Mandai, Takashi Shibuya, Hiroto Chaen, Shigeharu Fukuda, Toshiyuki Sugimoto, and Yoshio Tsujisaka*
Hayashibara Biochemical Laboratories, Inc., 7-7 Amase-minamimachi, Okayama
700, Japan
* Hayashibara Institute Corp., 1-2-3 Shimoishii, Okayama 700, Japan
Received December 26, 1996
@Cyclomaltodextrin glucanotransferase from Bacillus stearothermophilus
produced a series of glycosyl-trehaloses through the transglycosylation
reaction with cyclomaltohexaose as the glycosyl donor and trehalose as
its acceptor.
@After -amylase treatment, five species of glycosyl-trehaloses were
isolated by column chromatography. After chemical and enzymatic analyses,
it was concluded that these oligosaccharides were -maltosyl -D-glucopyranoside,
-maltotriosyl -D-glucopyranoside, -maltosyl -maltoside, -maltotriosyl
-maltoside, and -maltotriosyl -maltotrioside. These were not hydrolyzed
by salivary amylase, artificial gastric juice, or pancreatic amylase, however
they were hydrolyzed by enzymes of the small intestine.
Key words: trehalose; CGTase; transglycosylation
-19-
Effects of Medium-chain Fatty Acids and Their Acylglycerols
on the Transport of Penicillin V across Caco-2 Cell Monolayers
Motohiro Shima, Kaori Yohdoh, Masayo Yamaguchi, Yukitaka Kimura, Shuji Adachi, and Ryuichi Matsuno
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-01, Japan
Received December 26, 1996
@The transport-enhancing effects of medium-chain fatty acids (caproic,
caprylic, and capric acids) and their acylglycerols (mono-, di-, and triacylglycerols)
were investigated by using Caco-2 cell monolayers as a model of the human
intestinal epithelium. Penicillin V was used as a model for a hydrophilic
bioactive compound. Among the fatty acids and acylglycerols tested, 1,2-dicaproin,
monocaprin, monocaprylin, and capric acid sodium salt effectively enhanced
the transport rate, whereas other substances enhanced the rate only slightly
or not at all. With each of these four substances, the rate of enhancement
was proportional to the concentration at low concentrations, but leveled
off at high concentrations. The transport-enhancing effects were well correlated
with the reduction in surface tension and with a physico-chemical parameter,
denoted by the surface energy-lowering coefficient, characterizing the
surface activity of a substance.
Key words: Caco-2; transepithelial transport; medium-chain fatty acids
and acylglycerols; surface activity; physico-chemical property
-20-
Enhancing Effect of Interleukin-4 on the Secretion of
Interferon- by (/)s1-Casein-specific CD8+ T Cells
Ken-ichi Nishijima, Masako Kohyama, Tatsuhiro Hisatsune, Hiroko Kato, Masahiro Kakehi, and Shuichi Kaminogawa
Department of Applied Biological Chemistry, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113, Japan
Received January 8, 1997
@(/)s1-Casein-specific CD8+ T cell clones expressed the interleukin
(IL)-4 receptor, although they did not secrete detectable IL-4. We found
that IL-4 significantly enhanced the secretion of interferon (IFN)- by
these CD8+ T cell clones. IL-4 also enhanced the secretion of IFN- induced
by stimulating the immobilized anti-CD3 antibodies of polyclonal CD8+ T
cells which had been isolated from lymph nodes and were stimulated in vitro
with the immobilized anti-CD3 antibody and IL-2. In addition, IL-4 added
at the time of this first in vitro stimulation induced strong IFN- productivity,
as well as IL-4 and IL-10 productivity, which were detectable upon restimulation
of these cells. Results are discussed in relation to the inhibitory effects
of IFN- production on IL-4-producing cells.
Key words: interferon-; interleukin-4; CD8+ T cell; s1-casein
-21-
Further Studies on the Preparation of Low Sodium Chloride-containing
Soy Sauce by Using Ornithyltaurine Hydrochloride and Its Related Compounds
Rie Kuramitsu, Daisuke Segawa,* Kozo Nakamura,* Shunsuke Muramatsu,** and Hideo Okai*
Department of Chemistry, Faculty of General Education, Akashi College
of Technology, Uozumi, Akashi, Hyogo 674, Japan
* Department of Fermentation Technology, Faculty of Engineering, Hiroshima
University, 1-4-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 724, Japan
** Tenyo Takeda Co., Ltd., 9-30 Saiwai-cho, Kofu, Yamanashi 400, Japan
Received January 13, 1997
@Low sodium chloride-containing soy sauce samples were prepared by adding
salty peptide and its related compounds to pre-soy sauce. The soy sauce
obtained by adding Orn-TauEHCl was very similar to commercial soy sauce
in that it exhibited unadulterated saltiness and a good relative balance
between salty and umami tastes. The soy sauce obtained by adding Gly-OEtEHCl
exhibited a slightly stronger salty taste, while the one obtained by adding
LysEHCl had a slightly weaker salty taste at high concentrations than
commercial soy sauce. The soy sauce obtained with added KCl produced a
stronger umami taste at all concentrations than the commercial type. The
influence of ingredients other than NaCl and MSG in soy sauce (amino acids,
acids, sugars, and alcohol) on the salty and umami tastes of soy sauce
was also studied. In comparison with the NaCl substituents studied here,
these compounds were not as effective for influencing the apparent intensity
of the taste of soy sauce when they were individually added.
Key words: ornithyltaurine hydrochloride; salty peptide; low sodium chloride-containing
soy sauce
-22-
Purification and Characterization of Glutamate Decarboxylase
from Lactobacillus brevis IFO 12005
Yoshie Ueno, Kiyoshi Hayakawa, Saori Takahashi,* and Kohei Oda*,
Kyoto Prefectural Comprehensive Center for Small and Medium Enterprises,
17 Chudoji, Minamimachi, Shimogyo-ku, Kyoto 600, Japan
* Department of Applied Biology, Faculty of Textile Science, Kyoto Institute
of Technology, Mastugasaki, Sakyo-ku, Kyoto 606, Japan
Received January 17, 1997
@Glutamate decarboxylase (GAD) [EC 4.1.1.15] was purified from a cell-free
extract of Lactobacillus brevis IFO 12005 by chromatographies on Sephadex
G-100, DEAE-Sepharose CL-6B, and Mono Q. About 9 mg of purified GAD was
obtained from 90.2 g of wet cells. The purified preparation showed a single
protein band on SDS-PAGE. The molecular weights of purified GAD by SDS-PAGE
and gel filtration on Superdex 200 were 60,000 and 120,000, respectively,
indicating that GAD from L. brevis exists as a dimer. The N-terminal amino
acid sequence of the purified GAD was NH2-Met-Asn-Lys-Asn-Asp-Gln-Glu-Gln-Thr-.
The optimum pH and temperature of GAD were at pH 4.2 and at 30C. The
GAD activity was increased by the addition of sulfate ions in a dose-dependent
manner. The order of effect was as follows: ammonium sulfate > sodium
sulfate > magnesium sulfate, indicating that the increase of hydrophobic
interaction between subunits causes the increase of GAD activity. The purified
GAD reacted only with L-glutamic acid as a substrate and the Km, k(/)cat,
and k(/)cat/Km values were 9.3 mM, 6.5 s-1, and 7~102 M-1 s-1, respectively.
Key words: glutamate decarboxylase; gamma amino-n-butyric acid; Lactobacillus
brevis; purification; characterization
-23-
Isolation and Characterization of kar2-404 Mutation in
Saccharomyces cerevisiae
Akiko Kawamura-Watabe and Masao Tokunaga*,
Mitsubishi Kasei Institute of Life Sciences, 11 Minamiooya, Machida-shi,
Tokyo 194, Japan
* Laboratory of Applied Microbiology, Faculty of Agriculture, Kagoshima
University, 1-21-24 Korimoto, Kagoshima 890, Japan
Received January 27, 1997
@We have devised a direct screening method to isolate mutations in the
KAR2 gene, and have isolated a BiP/KAR2 mutant, kar2-404, from Saccharomyces
cerevisiae as a small halo-forming mutant of secreted mouse -amylase.
The mutation site was identified as a point mutation at t1337 to c1337
resulting in the Ile404Thr mutation of mature Kar2-404p, located at the
most NH2-terminal first -sheet structure (1) of the putative peptide-binding
domain. This isoleucine is highly conserved in the Hsp70 family. By pulse-chase
experiments, no obvious difference was detected in the intracellular secretion
rate of MF1-prepro-signal-mouse--amylase between the wild type and
the kar2-404 mutant. However, only about half the amount of secreted heterologous
protein, mouse -amylase, was detected in the mutant culture medium compared
with wild type. A smaller amount of homologous protein, -factor, was
also detected and decreased faster in the mutant culture medium than in
wild type. Kar2-404p was expressed about 3-fold more than wild type Kar2p,
probably to cover its defective functions, and the turnover rates of Kar2p
and Kar2-404p were about the same in vivo. The purified Kar2-404p was slightly
more sensitive to chymotryptic digestion than Kar2p in vitro.
Key words: KAR2; secretion; mouse -amylase; kar2-404 mutant
-24-
Separation and Determination of Yellow and Red Safflower
Pigments in Food by Capillary Electrophoresis
Toshiro Watanabe, Naoki Hasegawa, Akira Yamamoto, Shiro Nagai, and Shigeru Terabe*
Yaegaki Technology Development Laboratories, Yaegaki Bio-industry Inc.,
681 Hayashida, Himeji, Hyogo 679-42, Japan
* Faculty of Science, Himeji Institute of Technology, Kamigori, Hyogo 678-12,
Japan
Received February 10, 1997
@Capillary electrophoretic methods were developed for analyzing yellow
and red safflower pigments in food. Yellow safflower pigment was successfully
separated by capillary zone electrophoresis (CZE) with a 300 mM borate
buffer (pH 9.0), but this method could not successfully separate the red
safflower pigment. The red safflower pigment was discolored in an alkaline
solution. Both the yellow and red pigments could be successfully separated
by micellar electrokinetic chromatography (MEKC) with 2.0% butyl acrylate/butyl
methacrylate/methacrylic acid copolymer sodium salts (BBMA), but neither
pigment could be separated with 20 mM sodium dodecyl sulfate. The yellow
safflower pigment was extracted from food samples (juice and candies) by
solid-phase extraction cartridges and analyzed by the developed technique.
Key words: capillary zone electrophoresis; micellar electrokinetic chromatography;
safflower pigments
-25-
Note
Purification and Characterization of a Glycine-rich Polypeptide Tightly
Bound to Cell Walls from Soybean Aleurone Layers
Mitsuru Fukuda, Hiroko Noda, and Isao Toyosawa
Department of Food Science and Nutrition, Mukogawa Women's University, 6-46 Ikebiraki-cho, Nishinomiya, Hyogo 663, Japan
Received October 21, 1996
@A part of cell walls in soybean aleurone layers remained undigested after
pectinase and cellulase treatments, and the features of the undigested
cell walls were similar to those of Casparian strips. Glycine-rich polypeptides
(GRPP) were extracted with 0.4 N NaOH from the undigested cell walls, Casparian
strip-like tissues. Approximately 6.5-kDa GRPP obtained by gel-permeation
chromatography from the extract was purified by anion-exchange HPLC and
reverse-phase HPLC. The major amino acids of GRPP were glycine (69%) and
serine (13%). The N-terminal amino acid sequence of GRPP was the same polyglycine
as 30 kDa glycine-rich protein (GRP). GRPP would participate in adhesion
between neighboring cells in aleurone layers because of tight binding to
the cell wall.
Key words: glycine-rich polypeptide; aleurone layer; soybean; cell wall;
Casparian strip
-26-
Note
Characteristics of Escherichia coli HB101 and Pseudomonas putida PpY101
Harboring a Recombinant Plasmid with Tandem Insertion of the Mercury Resistance
Operon
Teruyo Kurabayashi,* Kazuhiro Iwasaki, Hiroo Uchiyama, Kunihiko Nakamura,** Hideo Tanaka,* and Osami Yagi
National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba,
Ibaraki 305, Japan
* Institute of Applied Biochemistry, University of Tsukuba, 1-1-1 Tennoudai,
Tsukuba, Ibaraki 305, Japan
** National Research Center for Minamata Disease, 4058-18 Hama, Minamata,
Kumamoto 867, Japan
Received October 28, 1996
@We constructed the plasmid pSUPmer2 by inserting tandem copies of the
mercury resistance (mer) operon into a broad host range-vector, and introduced
it into Escherichia coli HB101 and Pseudomonas putida PpY101 to increase
their mercury resistance. Strains harboring plasmid pSUPmer2 had higher
mercury resistance and mercuric reductase activity than those strains harboring
the plasmid pSUPmer which had one copy of the mer operon. Mercury resistance
of P. putida PpY101 was significantly increased by tandem insertion of
the mer operon.
Key words: mer operon; tandem gene amplification; mercury resistance; mercuric
reductase; bioremediation
-27-
Note
Distribution of Cu-resistant and Non-resistant Bacteria in Several Cu-contaminated
Soils in Japan: Usefulness of the Tolerance Level as an Index of Cu Contamination
in Soil
Takashi Kunito, Keishi Senoo, Kazunari Nagaoka, Hiroshi Oyaizu, and Satoshi Matsumoto
Department of Applied Biological Chemistry, Division of Agriculture and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113, Japan
Received November 15, 1996
@Seven Cu-contaminated soils were measured for the number of bacteria
grown on agar plates containing various concentrations of Cu, total Cu
contents, and DTPA-extractable Cu contents. The relationship between the
bacterial number grown on agar plates and concentration of Cu in the agar
plates (0-1.5 mM) was well fitted to Duxbury's exponential equation. Total
bacterial numbers in the soils positively correlated with the B values
in the equation. The B value seemed to be a better index of biotoxic Cu
amount in soil than DTPA-extractable Cu of soil. We offer a modification
of Duxbury's exponential equation, in which the B value can be estimated
once total soil bacterial number was counted.
Key words: heavy metals; copper; biomonitoring; tolerance level; pollution
-28-
Note
Effect of Chaotropic Salt on the Secondary Structure of Pigskin Gelatin
Yong Sik Cho and Kyung Bin Song
Department of Food Science and Technology, College of Agriculture, Chungnam National University, Taejon 305-764, Korea
Received November 15, 1996
@Pigskin gelatin was prepared and its molecular weight profile was examined
by SDS-PAGE. The major molecular weights of gelatin were 214 kDa, 135 kDa,
and 122 kDa. The secondary structure of a gelatin solution in the presence
of chaotropic salt was studied by using circular dichroism (CD). The CD
study clearly showed that the chaotropic salt increased the ordered secondary
structure of the gelatin solution due to the altered water structure.
Key words: pigskin gelatin; circular dichroism; secondary structure; chaotropic
salt
-29-
Note
Increase of Catalytic Activity of -Chymotrypsin by Metal Salts for Transesterification
of an Amino Acid Ester in Ethanol
Toshiya Sasaki and Hideo Kise
Institute of Materials Science, University of Tsukuba, 1-1-1 Tennoudai, Tsukuba, Ibaraki 305, Japan
Received December 10, 1996
@-Chymotrypsin-catalyzed transesterification of N-acetyl-L-tyrosine
methyl ester in ethanol was markedly accelerated by addition of small amounts
of divalent metal salts.
@The reaction rate depended not only on the nature of metal ions but also
on the nature of anionic counter ions. Calcium acetate was the most effective
among the metal salts used. The reaction followed Michaelis-Menten kinetics,
and it was found that the reaction increase is due to the increase in k(/)cat.
Key words: -chymotrypsin; transesterification; metal salt; ethanol
-30-
Note
Fluorescence-labeled Abscisic Acid Possessing Abscisic Acid-like Activity
in Barley Aleurone Protoplasts
Tadao Asami, Ling Tao,* Shin Yamamoto, Masumi Robertson,** Yong-Ki Min, Noboru Murofushi,* and Shigeo Yoshida
The Institute of Physical and Chemical Research (RIKEN ), 2-1 Hirosawa,
Wako, Saitama 351-01, Japan
* Department of Applied Biological Chemistry, The University of Tokyo,
Bunkyo-ku, Tokyo 113, Japan
** CSIRO Division of Plant Industry, GPO Box 1600, Canberra, ACT2601, Australia
Received December 19, 1996
@Novel fluorescence-labeled abscisic acid was synthesized by introducing
a fluorescent functional group into the 4' position of abscisic acid via
4-aminophenylcarbohydrazide as a spacer. The inhibitory activity toward
amylase induction by this chemical was evaluated by using aleurone protoplasts,
and it was demonstrated that the analogue possessed abscisic acid-like
activity.
Key words: abscisic acid; gibberellin; aleurone cells; protoplast; fluorescence
labeling
-31-
Note
Effects of Sex Hormones on the Metabolism of Tryptophan to Niacin and to
Serotonin in Male Rats
Katsumi Shibata and Satoko Toda
Department of Human Health Science, Faculty of Human Sciences, Osaka International University for Women, Moriguchi, Osaka 570, Japan
Received December 20, 1996
@It is known that deaths attributable to pellagra, which is considered
to be a disease caused by the disturbance of tryptophan metabolism, have
been approximately two-fold higher in women than in men. We investigated
the effects of the administration of female and male sex hormones on the
contents of tryptophan and such metabolites as serotonin, nicotinamide,
N 1-methylnicotinamide, N 1-methyl-2-pyridone-5-carboxamide, and N 1-methyl-4-pyridone-3-carboxamide,
and on the conversion ratio of tryptophan to niacin in male rats. Feeding
a diet containing estrone or testosterone had no effect on the concentrations
of tryptophan and serotonin in the blood and brain, or on the concentration
of 5-hydroxyindole-3-acetic acid in the brain. On the contrary, feeding
a diet containing estrone caused to a decrease in the urinary excretion
of nicotinamide, N 1-methylnicotinamide, N 1-methyl-2-pyridone-5-carboxamide,
and N 1-methyl-4-pyridone-3-carboxamide, and of the conversion ratio of
tryptophan to niacin when compared with the control rats. Feeding a diet
containing testosterone had no effect on any parameter. We postulate from
these findings that the cause of higher pellagra deaths in women than in
men is attributable to the decrease in the formation of niacin from tryptophan,
but not in the formation of serotonin by the female hormone. It seems likely
that female sex hormones inhibit the synthesis of niacin from tryptophan,
and that women, especially during pregnancy, will be more at risk to pellagra
than are men.
Key words: hormone; estrone; serotonin; niacin; tryptophan
-32-
Note
Chemical Characterization of a Sulfated Polysaccharide-Peptidoglycan Complex
from an Arthrobacter sp.
Ken-ichi Yamazaki,*, Makoto Suzuki,* Kazuyoshi Inukai,** and Hideo Hakusui **
* Basic Technology Research Laboratory and ** Drug Metabolism & Analytical Chemistry Research Laboratory, Daiichi Pharmaceutical Co., Ltd., 16-13 Kitakasai 1-chome, Edogawa-ku, Tokyo 134, Japan
Received December 27, 1996
@Structural features of the sulfated polysaccharide-peptidoglycan complex
from an Arthrobacter species were examined. The molecular weight was estimated
to be from 13,000 to 15,000. The complex mainly consists of galactose and
glucose at a molar ratio of 5 : 1. Glutamic acid, glycine, alanine, and
diaminopimelic acid (DAP) were found to be present at a molar ratio of
1 : 1 : 2 : 1.
Key words: cell wall; Arthrobacter; polysaccharide; peptidoglycan; MALDI-TOF
-33-
Note
Accelerating Effect of Chitosan Intake on Urinary Calcium Excretion by
Rats
Masahiro Wada, Yoshikazu Nishimura,* Yoshito Watanabe,* Toshichika Takita, and Satoshi Innami
Department of Nutrition, Tokyo University of Agriculture, 1-1-1 Sakuragaoka,
Setagaya-ku, Tokyo 156, Japan
* Division of Human Radiation Environment, National Institute of Radiological
Science, 4-9-1 Anagawa, Inage-ku, Chiba 263, Japan
Received January 16, 1997
@The effect of chitosan on calcium (47Ca) metabolism was investigated
in rats. The whole-body retention of 47Ca by rats fed on a 5% chitosan
diet was significantly decreased when compared with that of rats fed on
a cellulose diet, but showed no significant difference from that of rats
fed on a fiber-free diet. Although there was no significant difference
in the fecal excretion of 47Ca between the chitosan group and the cellulose
or fiber-free group, the urinary excretion of 47Ca was significantly increased
in the chitosan group when compared with the cellulose group. These results
suggest that dietary chitosan would affect the calcium metabolism in animals.
Key words: chitosan; calcium; rat
-34-
Note
Effect of Phosphoryl Oligosaccharides on Iron Solubility under Neutral
Conditions
Hiroshi Kamasaka, Kenji To-o, Kaname Kusaka, Takashi Kuriki, Takashi Kometani, and Shigetaka Okada
Biochemical Research Laboratory, Ezaki Glico Co., Ltd., 4-6-5 Utajima, Nishiyodogawa-ku, Osaka 555, Japan
Received January 23, 1997
@The solubility of iron ions with phosphoryl oligosaccharides (POs) from
a potato starch hydrolysate was investigated in the presence of bicarbonate
and phosphate salts. The oligosaccharides were separated into two fractions,
PO-1 and PO-2. PO-1, having one phosphoryl residue, solubilized equivalent
moles of iron, and PO-2, having two phosphoryl residues, solubilized more
than 10-fold equivalent moles of iron. The number of phosphate groups attached
to the molecule influenced the solubility of iron. The ability of PO-2
was equal to that of casein phosphopeptide (CPP) which is currently used
as an iron solubilizer for food.
Key words: phosphoryl oligosaccharides; potato starch; iron
-35-
Note
Construction of Escherichia coli-Bifidobacterium longum Shuttle Vector
Transforming B. longum 105-A and 108-A
Hajime Matsumura, Akio Takeuchi, and Yasunobu Kano
Department of Molecular Genetics, Institute of Molecular and Cellular Biology for Pharmaceutical Sciences, Kyoto Pharmaceutical University, 1 Shichono-cho, Misasagi, Yamashina-ku, Kyoto 607, Japan
Received January 30, 1997
@A shuttle vector, pBLES100, was constructed by cloning a Bifidobacterium
longum plasmid and a gene encoding spectinomycin adenyltransferase AAD(9)
from Enterococcus faecalis into the Escherichia coli vector pBR322. Stable
transformants with this plasmid were obtained with an efficiency of 2.2~104
transformants/g DNA or 6.9~10-5 transformants/cell/g DNA under the
optimal conditions of 10.0 kV/cm, 200 , and 25 F, using B. longum 105-A
harvested at late log phase of growth.
Key words: Bifidobacterium longum; Escherichia coli ; shuttle vector pBLES100;
transformation by electroporation; spectinomycin adenyltransferase AAD(9)
-36-
Note
Synthesis and Insecticidal Activity of New 2- and 6-Substituted 4-Acetoxyquinolines
Nobuto Minowa, Kei-ichi Imamura, and Seiji Shibahara
Pharmaceutical Research Center, Meiji Seika Kaisha, Ltd., 760 Morooka, Kohoku-ku, Yokohama 222, Japan
Received February 12, 1997
@The 4-acetoxyquinolines possessing 2-alkyl, alkenyl, and 6-fluoro groups
were synthesized and their insecticidal activities were evaluated.
Key words: 4-acetoxyquinolines; insecticidal activity
-37-
Note
A New Enzymatic Method for L-Phenylalanine Synthesis Using Aminoacylase
and Phenylalanine Dehydrogenase
Hong-Yon Cho and Kenji Soda*
Department of Food Science and Biotechnology, Graduate School of Biotechnology,
Korea University, Seoul 136-701, Korea
* Department of Biotechnology, Faculty of Engineering, Kansai University,
Suita, Osaka 564, Japan
Received February 24, 1997
@An aminoacylase, inducibly formed in Bacillus thermoglucosidius grown
with a synthetic compound, acetamidocinnamate, was used for enzymatic synthesis
of L-phenylalanine from chloroacetamidocinnamate. The reaction system consisted
of the hydrolysis of chloroacetamidocinnamate to phenylpyruvate by aminoacylase
and the reductive amination of phenylpyruvate to L-phenylalanine by phenylalanine
dehydrogenase. The coenzyme NADH consumed was regenerated by a coupled
reaction with formate dehydrogenase. Under optimum conditions for L-phenylalanine
production, more than 98% of the initially added chloroacetamidocinnamate
was converted effectively to L-phenylalanine without appreciable decomposition
or racemization.
Key words: aminoacylase; enzymatic synthesis; chloroacetamidocinnamate;
l-phenylalanine
-38-
Note
Solubilizing Coelenterazine in Water with Hydroxypropyl--cyclodextrin
Katsunori Teranishi and Osamu Shimomura
Marine Biological Laboratory, Woods Hole, MA 02543, U.S.A.
Received February 26, 1997
@Coelenterazine is poorly soluble in water except at alkaline pH values
that promote auto-oxidation. The solubility is markedly increased by adding
hydroxypropyl--cyclodextrin in a neutral buffer solution, the solubilization
being most effective in the pH range of 6.5 to 7.0. The concentration of
coelenterazine dissolved in a pH 7.0 buffer containing 50 mM hydroxypropyl--cyclodextrin
was about 280-fold higher than that without the additive.
Key words: coelenterazine; hydroxypropyl--cyclodextrin; solubility
-39-
Rapid Paper
Thiamine Increases Expression of Yeast Gene
Kimihisa Ichikawa, Yoichiro Shiba, Mitsuo Yamazaki, and Nobufusa Serizawa
Biomedical Research Laboratories, Sankyo Co., Ltd., 2-58 Hiromachi 1-chome, Shinagawa-ku, Tokyo 140, Japan
Received July 8, 1996
@We found that CPY production in Saccharomyces cerevisiae KS58-2D/pCY303
was increased by the addition of thiamine into the medium, while the addition
of thiamine had no effect on cell growth. It became clear that the positive
effect of thiamine was due to transcriptional increase, because the levels
of CPYmRNA were increased according to the amount of thiamine added. Furthermore,
it was suggested that thiamine generally increases the expression of yeast
genes, since the expression of the luciferase gene that was artificially
constructed was also increased to some extent by thiamine in S. cerevisiae.
Key words: thiamine; carboxypeptidase Y; Saccharomyces cerevisiae; transcriptional
activation
-40-
Rapid Paper
An Anti-hydrotactic Response and Solid Surface Recognition of Germ Tubes
of the Rice Blast Fungus, Magnaporthe grisea
Jin-zhong Xiao, Tadakazu Watanabe, Shigeko Sekido, Woo-Bong Choi, Takashi Kamakura, and Isamu Yamaguchi
Microbial Toxicology Laboratory, The Institute of Physical and Chemical Research (RIKEN ), Wako, Saitama 351-01, Japan
Received March 3, 1997
@Magnaporthe grisea, the causal agent of rice blast disease, differentiates
an appressorium to penetrate through the host cuticle with an infection
peg elaborated from the appressorium. Similar reaction is observed on various
synthetic substrata. By using a hardness-adjustable material, Hycel A-342,
the correlation between appressorium formation and substratum hardness
was evaluated. The results suggested that substratum hardness played an
important role in triggering appressorium formation of M. grisea. Furthermore,
growth orientation of germ tubes was coerced by the hydrophobicity of the
contact surfaces. When conidia germinated at the interface of two differently
hydrophobic phases, the germ tubes grew preferentially towards the more
hydrophobic phase. Mutagenesis studies suggested that loss of this anti-hydrotactic
behavior impaired appressorium formation. Since water is a pre-requisite
for germination, we propose that the anti-hydrotactic response initiates
attempted penetration into the plants by germ tubes, then triggers appressorium
formation by the surface hardness.
Key words: Magnaporthe grisea; Pyricularia oryzae; appressorium formation;
surface recognition; plant pathogenicity
-41-
Rapid Paper
Conformational Behavior of Chitosan in the Acetate Salt: An X-Ray Study
Atsushi Yamamoto, Junpei Kawada, Toshifumi Yui,* and Kozo Ogawa
Research Institute for Advanced Science and Technology (RIAST ), Osaka
Prefeature University, 1-2 Gakuen-cho, Sakai, Osaka 593, Japan
* Faculty of Engineering, Miyazaki University, Miyazaki 889-21, Japan
Received March 3, 1997
@A well-defined X-ray fiber pattern of chitosan acetate was obtained by
immersing a tendon chitosan, prepared from a crab tendon chitin by a solid-state
N-deacetylation, in an aqueous acetic acid-isopropanol solution at 110C.
This pattern was very similar to that of chitosan salts with some inorganic
acids, such as HF, HCl, and H2SO4, in which chitosan chains form an 8/5
helix, indicating that chitosan acetate also take up this conformation.
This information may give an influential clue to the chitosan conformation
in the aqueous acetic acid solution, the most popular solvent for chitosan.
However, after one month of storage of the chitosan acetate, the fiber
pattern, the density and its IR spectrum changed to those of the anhydrous
polymorph of chitosan, suggesting that the acetic acid was removed accompanied
with water molecules from the crystal during storage and that the polymorph
can be obtained not only by annealing chitosan, but also through the chitosan
acetate.
Key words: X-ray fiber pattern; chitosan acetate; 8/5 helix; anhydrous
chitosan