(Vol.62 No.1 1998)
Effects of Orally Ingested Bifidobacterium longum on the
Mucosal IgA Response of Mice to Dietary Antigens
Takeshi TAKAHASHI, Emiko NAKAGAWA, Toshi NARA, Takaji YAJIMA, and Tamotsu
KUWATA 10
Casein Phosphopeptides from Casein Micelles by Successive
Digestion With Pepsin and Trypsin
Tomotada ONO,Yasushi TAKAGI,Isao KUNISHI 16
Iron Chelation by Chlorogenic Acid as a Natural Antioxidant
Yasuhisa KONO, Sakiko KASHINE, Takushi YONEYAMA, Yuji SAKAMOTO, Yoshihisa
MATSUI, and Hitoshi SHIBATA 22
A Cysteine Protease from Young Stems of Asparagus: Isolation,
Properties, and Substrate Specificity
Hiroo YONEZAWA, Makoto KANEDA, and Tetsuya UCHIKOBA 28
Osteopathy in Broiler Chicks Fed Toxic Mimosine in Leucaena
leucocephala
Yasuhiro KAMADA, Nobuaki OSHIRO, Mayumi MIYAGI, Hirosuke OKU, Fujiya
HONGO,* and Isao CHINEN 34
Purification and Some Properties of p-Nitrophenyl--D-glucoside-hydrolyzing
Enzymes in Culture Filtrate of Bacillus circulans KA-304 Grown on Cell-wall
Preparation of Schizophyllum commune
Katsushige MIZUNO, Naoyuki AWAZU, and Takashi TACHIKI 39
Effects of Japanese Black Tea on Atherosclerotic Disorders
Takako YOKOZAWA, Erbo DONG, Takako NAKAGAWA,* Dong Wook KIM, Masao HATTORI,
and Hitomi NAKAGAWA* 44
Role of Sucrose in Gamma-irradiated Chrysanthemum Cut Flowers
Kazuhiko NAKAHARA, Olivia Kimiko KIKUCHI, Setsuko TODORIKI, Hiroshi HOSODA,
and Toru HAYASHI 49
Distribution of Defense-Related Enzymatic Activities in
the Quiescent Organs of Phaseolus vulgaris L. Seeds
Ana Luiza RAMOS, Guillermo SEIJO, Chiharu ISSHIKI, and Tohru IWAMOTO 54
Cloning and Characterization of a Chitinase-encoding Gene
(chiA) from Aspergillus nidulans, Disruption of Which Decreases Germination
Frequency and Hyphal Growth
Naoki TAKAYA, Daisuke YAMAZAKI, Hiroyuki HORIUCHI, Akinori OHTA, and Masamichi
TAKAGI 60
Gene Cloning and Characterization of Thermostable Lipase
from Bacillus stearothermophilus L1
Hyung-Kwoun KIM, Sun-Yang PARK, Jung-Kee LEE, and Tae-Kwang OH 66
Synthesis of Some 1-Methyladenine Analogs and Their Biological
Activities on Starfish Oocyte Maturation
Tetsuo TORAYA, Tetsuo KIDA, Sei-ichi TANAKA, Masaki MATSUSHITA, Taro TSURUKAI,
and Hidenori SHIOTSUKA 72
Molecular Interaction between Proteins Involved in EvgAS
Signal Transduction of Escherichia coli
Hiroyuki TANABE, Takayuki MASUDA, Katsuhide YAMASAKI, Akinori KATOH,
Sachiko YOSHIOKA, and Ryutaro UTSUMI 78
The Relationship between Transport-enhancement Effects
and Cell Viability by Capric Acid Sodium Salt, Monocaprin, and Dicaproin
Motohiro SHIMA, Yukitaka KIMURA, Shuji ADACHI, and Ryuichi MATSUNO 83
Primary Structure of a Squid Acid and Base Non-specific
Ribonuclease
Ayumu KUSANO, Masanori IWAMA, Kazuko OHGI, and Masachika IRIE 87
Phenylphenalenone-type Phytoalexins from Unripe BuEngulan
Banana Fruit
Tsunashi KAMO,1,2 Nagomi KATO,1 Nobuhiro HIRAI,1,2, Mitsuya
TSUDA,3 Daie FUJIOKA,4 and Hajime OHIGASHI1 95
Near Infra Red Detection of Internally Moldy Nuts
Susumu HIRANO, Noriyuki OKAWARA, and Shuichirou NARAZAKI 102
Potassium Iodide-Induced Changes in Pyruvate Dehydrogenase
Complex from Bacillus stearothermophilus
Yoichi ASO, Yasuaki HIROMASA, Yoshikatsu AIKAWA, Kohji MENO, and Masatsune
ISHIGURO 108
Purification and Characterization of an Isomaltotriose-producing
Endo-dextranase from a Fusarium sp.
Eishun SHIMIZU, Takehiro UNNO,* Manabu OHBA,** and Gentaro OKADA**,
117
Glucan Binding Regions of Dextransucrase from Leuconostoc
mesenteroides NRRL B-512F
Kazumi FUNANE, Tetsuya OOKURA, and Mikihiko KOBAYASHI 123
Chitin Binding Protein (CBP21) in the Culture Supernatant
of Serratia marcescens 2170
Kazushi SUZUKI,1 Megumi SUZUKI,1 Mayumi TAIYOJI,1 Naoki NIKAIDOU,1,2
and Takeshi WATANABE1,2 128
Expression of Biologically Active Human Calpastatin in
Baculovirus-infected Insect Cells and in Escherichia coli
Kiyotaka HITOMI, Akiko YOKOYAMA, and Masatoshi MAKI 136
Note
Effects of Bay m 1099, an a-Glucosidase Inhibitor, on Starch Degradation
in Germinating Mung Beans
Yotaro KONISHI, Michio AITANI, and Nobuji NAKATANI 142
Silk Protein, Sericin, Inhibits Lipid Peroxidation and
Tyrosinase Activity1
Norihisa KATO,*,2 Seiji SATO,* Atsushi YAMANAKA,* Hideyuki YAMADA,**
Naozumi FUWA,** and Masakazu NOMURA** 145
Note
Molecular Cloning and Characterization of a cDNA Encoding Asparagine Synthetase
from Soybean (Glycine max L.) Cell Cultures
Hiroshi YAMAGATA,*, Ayumi NAKAJIMA,* Chris BOWLER,** and Teruo IWASAKI*
148
Note
Increase of Foreign Gene Expression in Monocot and Dicot Cells by an Intron
in the 5? Untranslated Region of a Soybean Phosphoenolpyruvate Carboxylase
Gene
Tomohiko KATO, Emiko ITOH, Robert F. WHITTIER, and Daisuke SHIBATA 151
Note
Purification and Characterization of the 2-Ketoaldonate Reductase from
Brevibacterirnm ketosoreductum ATCC21914
Do-Young YUM, Sung-Sook BAE, and Jae-Gu PAN 154
Note
Effect of Antioxidants in Preventing the Thermal Decomposition of Phosphatidylcholine
Hydroperoxide
Naomichi BABA, Ikue TAKAHASHI, Hiroko DAIDO, Md. Khorshed ALAM, Shuhei
NAKAJIMA, and Takao KANEKO* 157
Note
Analysis of Glycerophospholipid Hydroperoxides by Ion Spray Mass Spectrometry
Naomichi BABA, Hiroko DAIDO, Tomoko KOSUGI, Miwako MIYAKE, and Shuhei NAKAJIMA
160
Note
Effects of Unsaturated Uronic Acid Residues at Non-reducing End on Bond
Cleavage Frequency of Poly(1,4--L-guluronide) Lyase from Enterobacter
cloacae M-1
Tomoko SHIMOKAWA, Shigeki YOSHIDA, and Isao KUSAKABE 164
Note
Escherichia coli Transformant Expressing the Glucose Dehydrogenase Gene
from Bacillus megaterium as a Cofactor Regenerator in a Chiral Alcohol
Production System
Michihiko KATAOKA, Luh Poni Sri ROHANI, Masaru WADA, Keiko KITA,*
Hideshi YANASE,* Itaru URABE,** and Sakayu SHIMIZU 167
Note
Effects of Substrate Solubility in Interesterification with Triolein by
Immobilized Lipase in Supercritical Carbon Dioxide
Seung-Heon YOON,1,2 Hideki NAKAYA,1 Osamu ITO,1 Osato MIYAWAKI,1
Kwan-Hwa PARK,2 and Kozo NAKAMURA1 170
Note
Pyricuol, a New Phytotoxin from Magnaporthe grisea
Jin-Cheol KIM, Ji-Young MIN, Heung-Tae KIM, Kwang-Yun CHO, and Seung-Hun
YU* 173
Note
Evaluation of the Antioxidative Activity of Tea by an Oxygen Electrode
Method
Midori KUMAMOTO and Tamiyoshi SONDA 175
Note
Inhibition of Glucan Synthesis by Casein Polymers Crosslinked by Glutaraldehyde
Osamu HAYASHIDA, Fumiki MORI, Keiji HASUMI, and Akira ENDO 178
Note
Reduction of Alkyl (2-Oxocyclohexyl)acetates by Baker's Yeast
Makoto GANAHA, Yuhei FUNABIKI, Minoru MOTOKI, Satoshi YAMAUCHI, and Yoshiro
KINOSHITA 181
Note
The Characterization of Acetic Acid Bacteria Efficiently Producing Bacterial
Cellulose from Sucrose: The Proposal of Acetobacter xylinum subsp. nonacetoxidans
subsp. Nov.
Yukiko KOJIMA, Naoto TONOUCHI, Takayasu TSUCHIDA, Fumihiro YOSHINAGA, and
Yuzo YAMADA 185
Note
Simple Method for Detecting Glycoproteins Dot-blotted or Electro-blotted
on to a Polyvinylidene Difluoride Membrane
Shizuya KABUTO and Koji MURAMOTO 188
Note
Novel Stereoselective Reaction of Levoglucosenone with Furfural
Toshio NISHIKAWA, Hiroshi ARAKI, and Minoru ISOBE* 190
Short Communication
Compensation for D-Glutamate Auxotrophy of Escherichia coli WM335 by D-Amino
Acid Aminotransferase Gene and Regulation of murI Expression
Lidong LIU,1 Tohru YOSHIMURA,1 Keiji ENDO,1 Kazuhisa KISHIMOTO,1
Yoshihiro FUCHIKAMI,1 James M. MANNING,2 Nobuyoshi ESAKI,1
and Kenji SODA3 193
-1-
Review
Biochemical Synthesis of Several Chiral Insecticide Intermediates and Mechanisms
of Action of Relevant Enzymes
Hideo HIROHARA* and Masako NISHIZAWA**
*Department of Materials Science, The University of Shiga Prefecture,
Hassaka, Hikone 522, Japan
**Biotechnology Laboratory, Sumitomo Chemical Co., Takatsukasa, Takarazuka,
Hyogo 665, Japan
Efficient biochemical processes were developed for the synthesis of several chiral alcohol and acid intermediates of insecticides by a combination of strictly stereoselective hydrolytic enzyme-catalyzed reactions and subsequent chemical transformations with inversion or racemization of the chiral center of the undesired antipodes. The whole amounts of starting racemic mixtures are converted to desired stereoisomers in the processes, which are generally applicable to the industrial productions of various chiral secondary alcohols and a-substituted acids once a highly stereospecific enzyme is obtained for the target compounds. The alcohols reported here are: 1-(4-phenoxyphenoxy)-2-propanol, 1; 4-hydroxy-3-methyl-2-(2'-propynyl)-2-cyclopentenone, 2; and a-cyano-3-phenoxybenzyl alcohol, 3. The acids are 2, 2-dimethyl-3-(2-methyl-1-propenyl)-cyclopropanecarboxylic acid (chrysanthemic acid), 4; and 2-(4-chlorophenyl)-3-methylbutyric acid, 5. In addition, the mechanism of action of Pseudomonas cepacia lipase (PCL), the most effective enzyme for the resolution of 1, and the recombinant Arthrobacter globiformis esterase (AES) for 4 was studied from the reaction kinetics. The site-directed mutagenesis techniques were also used for AES. The results indicated that the stereoselectivity of PCL is caused by the position and direction of a medium-sized substituent at the chiral center of the alcohol moiety in the rate-determining breakdown of a tetrahedral intermediate in the acylation of the enzyme and that the catalytic site of AES has the characteristics of the penicillin-recognizing enzymes in which Ser 59 in the concensus motif Ser-X-X-Lys plays a vital role as a nucleophile during acylation and Lys 62 acts as a general base.
-2-
Effects of Orally Ingested Bifidobacterium longum on the
Mucosal IgA Response of Mice to Dietary Antigens
Takeshi TAKAHASHI, Emiko NAKAGAWA, Toshi NARA, Takaji YAJIMA, and Tamotsu KUWATA
Nutrition Science Institute, Meiji Milk Products Co., Ltd., 540 Naruda, Odawara 250, Japan
Received April 16, 1997
To study the effects of lactic acid bacteria on the mucosal defence against
dietary protein antigens, we compared the mucosal IgA responses to -lactoglobulin
(-LG) of two groups of mice fed a whey protein diet with and without
a culture condensate of Bifidobacterium longum. Both total IgA and anti--LG
IgA levels in tissue extracts of the small intestinal wall were significantly
higher in mice fed the B. longum diet for 2 weeks than in control ones.
Peyer's patch (PP) cells from B. longum-fed mice had a much larger increase
in in vitro IgA production than ones from control mice. Furthermore, the
in vitro IgA response to -LG was detected only when PP cells from B.
longum-fed mice were assayed. These results suggest that orally ingested
lactic acid bacteria may protect a host from invasion of the intestinal
mucosa by dietary antigens that have escaped enzymatic digestion in the
intestine.
-3-
Casein Phosphopeptides from Casein Micelles by Successive
Digestion with Pepsin and Trypsin
Tomotada ONO, Yasushi TAKAGI, and Isao KUNISHI
Department of Bioscience and Technology, Iwate, University, Morioka 020, Japan
Received April 21, 1997
When milk is ingested, casein micelles will be successively digested by
pepsin in the stomach and trypsin in the intestine. Therefore, we digested
casein micelles successively with pepsin at pH 4.0 and trypsin at pH 7.0,
and recovered casein phosphopeptides (CPP) as CPP-calcium phosphate (CP)
complexes. The CPP-CP complexes contained 248 mg of calciumEg peptides
and 175 mg of inorganic phosphorusEg peptides, which were higher than
those of CPP-CP complexes driven from acid-precipitated casein and casein
micelles by tryptic digestion only. It contained more aS1-CN-5P(f59-79)
than the other CPP preparations did.
-4-
Iron Chelation by Chlorogenic Acid as a Natural Antioxidant
Yasuhisa KONO, Sakiko KASHINE, Takushi YONEYAMA, Yuji SAKAMOTO, Yoshihisa MATSUI, and Hitoshi SHIBATA
Department of Life Science and Biotechnology Faculty of Life and Environmental Science Shimane University, Matsue, Shimane 690, Japan
Received April 28, 1997
Chlorogenic acid, a dietary antioxidant, effectively inhibited the iron-induced
lipid peroxidation of bovine liver microsomes in a concentration-dependent
manner. In the Fenton-type reaction, chlorogenic acid inhibited the production
of the hydroxyl radical by iron-EDTA or iron-ADP, while iron plus chlorogenic
acid did not generate the hydroxyl radical. The formation of an iron complex
with chlorogenic acid was demonstrated by UVEvis absorbance spectroscopic,
ESR and 1H-NMR studies. The ferric complex with chlorogenic acid was
in the ferric high-spin state near rhombicity, and had no radical scavenging
activity. The results indicate that chlorogenic acid prevented the formation
of the hydroxyl radical by forming a chelate with iron whose complex cannot
catalyze the Fenton-type reaction.
-5-
A Cysteine Protease from Young Stems of Asparagus: Isolation,
Properties, and Substrate Specificity
Hiroo YONEZAWA, Makoto KANEDA, and Tetsuya UCHIKOBA
Laboratory of Biochemistry, Department of Chemistry, Faculty of Science, Kagoshima University, 1-21-35, Korimoto, Kagoshima 890, Japan
Received April 30, 1997
A protease was purified from the growing point of asparagus, Asparagus
officinalis, using a cystatin-Sepharose column. The asparagus protease
is the first protease isolated from the growing point of a plant tissue
and from Liliaceae. Its molecular mass was estimated to be 28 kDa by SDS-PAGE.
The optimum pH of the enzyme was 7 at 30EC using casein as a substrate.
The enzyme was strongly inhibited by monoiodoacetic acid, but not by diisopropylfluorophosphate,
suggesting that it is a cysteine protease. Asparagus protease had broad
specificity on the hydrolysis with oxidized B-chain of insulin as a substrate.
However, for the P2 position of the cleavage site, the large hydrophobic
side chains of amino acid residues such as Phe, Val, and Leu were considerably
preferred. Asparagus protease was similar to papain about specificity at
the P2 position. The N-terminal sequence of the first 12 residues was
identified and 8 residues among them agreed with that of papain, accompanying
the addition of one residue (Ala) to that of papain.
-6-
Osteopathy in Broiler Chicks Fed Toxic Mimosine in Leucaena
leucocephala
Yasuhiro KAMADA, Nobuaki OSHIRO, Mayumi MIYAGI, Hirosuke OKU, Fujiya HONGO,* and Isao CHINEN
Laboratories of Applied Biochemistry,
*Applied Bioresource Science, Faculty of Agriculture, University of The
Ryukyus, Nishihara, Okinawa 901-01, Japan
Received May 19, 1997 @
@Further studies of mimosine toxicity in broiler chicks were done to clarify
a possibility of osteopathy.
@The mineral content and density of femur and the strength, ductility,
and toughness for the index of mechanical properties significantly decreased
in the 1% mimosine group, compared with those in the control and restricted
groups. The stiffness had a decreasing tendency in the 1% mimosine group.
Consequently, it was concluded that chicks fed ad libitum a 1% mimosine
diet for 12 days developed osteopathy.
@The bone mineral density and the strength of the restricted group were
lower than those of the control group, and those of the 1% mimosine group
were still lower than those of the restricted group. Contents of pyridinoline
and deoxypyridinoline in the excrement were significantly higher in the
restricted group than those in the control group, but the contents in the
1% mimosine group were significantly lowest among the groups. Osteopathy
in chicks fed mimosine, therefore, seemed to be done by loss of appetite
and changing to a low turnover of bone caused by mimosine.
-7-
Purification and Some Properties of p-Nitrophenyl--D-glucoside-hydrolyzing
Enzymes in Culture Filtrate of Bacillus circulans KA-304 Grown on Cell-wall
Preparation of Schizophyllum commune
Katsushige MIZUNO, Naoyuki AWAZU, and Takashi TACHIKI
Department of Bioscience and Biotechnology, Faculty of Science and Engineering, Ritsumeikan University, Noji Higashi 1-1-1, Kusatsu 525-77, Japan
Received May 29, 1997
Hydrolyzing activities toward p-nitrophenyl (p-NP)--D-glucoside and laminarin
in a culture filtrate of Bacillus circulans KA-304, which has been observed
to form protoplasts from Schizophyllum commune mycelia, increased when
the bacterium was grown on a cell-wall preparation (CWP) of S. commune
or laminarin as a carbon source. An analysis of the filtrate with the CWP
suggested occurrence of two major p-NP--D-glucoside-hydrolyzing enzymes
(-D-glucosidases I and II) and a laminarin-hydrolyzing enzyme. After
separation by DEAE-cellulose column chromatography, -D-glucosidases I
and II were isolated (-D-glucosidase I: 13-fold purification with 34E
yield; -D-glucosidase II: 26-fold with 8E). The enzymes resembled each
other in their properties except for their molecular weight, subunit structure
(-D-glucosidase I: 200,000, tetramer; II: 100,000, dimer), and susceptibility
to such substances as p-chloromercuribenzoic acid and Ag{ ion. -D-Glucosidases
I and II hydrolyzed gentiobiose (-1,6 glucosidic linkage; Km3.6 mM,
-D-glucosidase I; 4.6 mM, -D-glucosidase II) and laminaribiose (-1,3
glucosidic linkage; Km6.1 mM, -D-glucosidase I; 6.7 mD -D-glucosidase
II), and showed a certain reactivity toward laminarin as well.
-8-
Effects of Japanese Black Tea on Atherosclerotic Disorders
Takako YOKOZAWA, Erbo DONG, Takako NAKAGAWA,* Dong Wook KIM,
Masao HATTORI, and Hitomi NAKAGAWA*
Research Institute for Wakan-Yaku, Toyama Medical and Pharmaceutical
University, Sugitani, Toyama 930-01, Japan
*Faculty of Education, Toyama University, Gofuku, Toyama 930, Japan
Received June 11, 1997
The atherogenic index was found to be significantly better in rats fed
a high-cholesterol diet supplemented with black tea extract than in the
ones not given the extract. It was also evident that black tea inhibited
the proliferation of smooth muscle cells involved in the development and
progression of atherosclerosis, and suppressed the production of oxidized
low-density lipoprotein, a cause of lipid accumulation. It thus seems likely
that black tea has an antiatherosclerotic action.
-9-
Role of Sucrose in Gamma-irradiated Chrysanthemum Cut Flowers
Kazuhiko NAKAHARA, Olivia Kimiko KIKUCHI, Setsuko TODORIKI, Hiroshi HOSODA, and Toru HAYASHI
National Food Research Institute, Ministry of Agriculture, Forestry and Fisheries, Kannondai, Tsukuba, Ibaraki 305, Japan
Received June 11, 1977
Vase solution containing 2% sucrose prevented the deterioration of chrysanthemum
(Dendranthema grandiflorum Kitamura) cut flowers induced by gamma-rays
at 750 Gy. Glucose, fructose, and sucrose in florets and leaves of irradiated
chrysanthemums decreased more rapidly than those of unirradiated ones,
when the cut chrysanthemums were held in a vase solution without sucrose.
The sugar contents of florets and leaves and the respiratory rate of irradiated
chrysanthemums held with sucrose remained at higher levels than those of
unirradiated ones. Incorporation of 14C from [14C]sucrose into
CO2 was increased by irradiation. Incorporation of [a-32P]dTTP
into trichloroacetic acid (TCA) insoluble substances in florets was increased
by irradiation and by exogenous sucrose supply. These results suggest that
sucrose in a vase solution was used as a respiratory substrate and facilitated
the repair of radiation-induced damage, resulting in the extension of longevity
of irradiated chrysanthemums.
-10-
Distribution of Defense-Related Enzymatic Activities in
the Quiescent Organs of Phaseolus vulgaris L. Seeds
Ana Luiza RAMOS, Guillermo SEIJO, Chiharu ISSHIKI, and Tohru IWAMOTO
Faculty of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama 790, Ehime, Japan
Received June 20, 1997
Analysis of quiescent seed extracts of Phaseolus vulgaris cultivars found
high activities of chitinase, N,N?-diacetylchitobiase and -N-acetylhexosaminidase
in whole seeds and their dissected organs (cotyledons, axis, and seed coat).
Activities of these enzymes were compared in seeds of two cultivars phenotypically
distinguishable by soft white (cv. Surattowonder) and hard brown (cv. Maisugata)
seed coats. In both cultivars, chitinase activity was found high in all
organs, chitobiase in the seed coat, and -N-acetylhexosaminidase mainly
in the axis. In terms of specific activity, all three enzymes were extraordinarily
higher in extracts of seed coat than the others, specially referring to
the cultivar Surattowonder. Although the cultivars showed in general similar
distribution patterns of activities among their seed organs, the discrepancies
found between them seem to be expressing intrinsic attributes of their
seed coats. The relationship between the two cultivars, the enzyme activities
measured and defense mechanisms are discussed.
-11-
Cloning and Characterization of a Chitinase-encoding Gene
(chiA) from Aspergillus nidulans, Disruption of Which Decreases Germination
Frequency and Hyphal Growth
Naoki TAKAYA, Daisuke YAMAZAKI, Hiroyuki HORIUCHI, Akinori OHTA, and Masamichi TAKAGI
Department of Biotechnology, The University of Tokyo, Yayoi, Bunkyo-ku, Tokyo 113, Japan
Received June 23, 1997
We cloned a chitinase-encoding gene from Aspergillus nidulans by polymerase
chain reaction using degenerated oligonucleotide primers designed from
the conserved amino acid sequences among chitinases from yeasts and Rhizopus
spp. The cloned gene, named chiA, encoded a polypeptide consisting of 660
amino acids. Disruption of chiA had no effect on hyphal or conidiophore
morphology, but germination frequency and hyphal growth rate decreased
substantially. Expression of chiA was investigated using Escherichia coli
-galactosidase as a reporter enzyme. The -galactosidase activity was
present during hyphal growth and increased twice as the conidiophores developed.
In situ staining of -galactosidase activity found high expression in
metulae, phialides, and conidia during conidiophore development, indicating
that the expression of chiA is developmentally regulated. This is the first
report to isolate a chitinase gene from A. nidulans and investigate its
functions using the gene disruption technique and gene fusion methods in
filamentous fungi.
-12-
Gene Cloning and Characterization of Thermostable Lipase
from Bacillus stearothermophilus L1
Hyung-Kwoun KIM, Sun-Yang PARK, Jung-Kee LEE, and Tae-Kwang OH
Microbial Enzyme RU, Korea Research Institute of Bioscience & Biotechnology (KRIBB), P.O. Box 115, Yusong, Taejon, 305-600, Korea
Received July 3, 1997
The gene coding for an extracellular lipase of Bacillus stearothermophilus
L1 was cloned in Escherichia coli. Sequence analysis showed an open reading
frame of 1254 bp, which encodes a polypeptide of 417 amino acid residues.
The polypeptide was composed of a signal sequence (29 amino acids) and
a mature protein of 388 amino acids. An alanine replaces the first glycine
in the conserved pentapeptide (Gly-X-Ser-X-Gly) around the active site
serine. The expressed lipase was purified by hydrophobic interaction and
ion exchange chromatography using buffers containing 0.02% (vEv) Triton
X-100. The lipase was most active at 60-65EC and in alkaline conditions
around pH 9-10. The lipase had highest activity toward p-nitrophenyl caprylate
among the synthetic substrates and tripropionin among the triglycerides.
It hydrolyzed beef tallow and palm oil more rapidly than olive oil at 50EC.
-13-
Synthesis of Some 1-Methyladenine Analogs and Their Biological
Activities on Starfish Oocyte Maturation
Tetsuo TORAYA, Tetsuo KIDA, Sei-ichi TANAKA, Masaki MATSUSHITA, Taro TSURUKAI, and Hidenori SHIOTSUKA
Department of Bioscience and Biotechnology, Faculty of Engineering, Okayama University, Tsushima-Naka, Okayama 700, Japan
Received July 4, 1997
Starfish oocytes are naturally arrested at the prophase stage of the first
meiotic division and resume meiosis in response to the maturation-inducing
hormone 1-methyladenine. Five analogs of 1-methyladenine including three
novel ones were synthesized and tested for biological activities as 1-methyladenine
agonists or antagonists in triggering reinitiation of meiosis of starfish
Asterina pectinifera oocytes, as well as for competition in binding to
putative 1-methyladenine receptors with respect to 1-methyladenine. 1-Ethyladenine
was an effective agonist, but 1-propyladenine served as a weak antagonist
to 1-methyladenine, indicating strict specificity for a relatively small
N-1 substituent. Analogs in which carboxymethyl or methyl group substitutes
for a hydrogen of 6-amino group still retained oocyte maturation-inducing
activity, but to a much lesser degree. The results of the competitive binding
assay with cortices of oocytes demonstrated that these agonists or antagonist
inhibited the binding of [3H]1-methyladenine to receptors. 8-methylamino-1-methyladenine
competed only weakly with [3H]1-methyladenine for the binding to cortices,
although it behaved as a potent antagonist.
-14-
Molecular Interaction between Proteins Involved in EvgAS
Signal Transduction of Escherichia coli
Hiroyuki TANABE, Takayuki MASUDA, Katsuhide YAMASAKI, Akinori KATOH, Sachiko YOSHIOKA, and Ryutaro UTSUMI
Department of Agricultural Chemistry, Kinki University, Naka-machi, Nara 631, Japan
Received July 7, 1997
EvgA and EvgS constitute one two-component signal transduction system in
Escherichia coli. Although probable signaling domains of these proteins
have been estimated, the molecular mechanism of their inteaction remains
to be elucidated. Here, we investigated protein to protein interactions
between EvgA and EvgS and also between the EvgAS system and other related
signaling pathways by means of surface plasmon resonance. EvgA and EvgS
interacted directly and inhibition of phosphorylation of their functional
domains abolished formation of the EvgAS complex. No interaction was observed
either between EvgA and Bordetella BvgS or BvgA and EvgS. OmpR, a response
regulator for the osmoregulative gene expression of E. coli, had similar
but not identical behavior towards EvgS to that of EvgA. These results
indicate that interaction between the signaling proteins is closely related
to phosphorylation of the functional domain of the proteins.
-15-
The Relationship between Transport-enhancement Effects
and Cell Viability by Capric Acid Sodium Salt, Monocaprin, and Dicaproin
Motohiro SHIMA, Yukitaka KIMURA, Shuji ADACHI, and Ryuichi MATSUNO
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto 606-01, Japan
Received July 7, 1997
@Using Caco-2 cell monolayers and MTT assay, the relationship between
cell viability (a) and transport-enhancement effect of 1,2-dicaproin (C6DG),
monocaprin (C10MG), and capric acid sodium salt (C10FANa) was examined.
Transport enhancement effect was assessed by apparent permeability (Papp)
of penicillin V. There was a linear relationship between (Papp|aSa)
and (1|a) values, where Sa was the apparent permeability for the viable
cells. The apparent permeability for the damaged cells (Sd) was evaluated
from the slope of the line. Each of the enhancer compounds gave a different
Sd value 2.00~10|4, 0.82~10|4, and 0.10~10|4 cmEs
for C6DG, C10MG, and C10FANa, respectively, but the value was independent
of its concentration for C10MG and C10FANa. C6DG would be the safest enhancer
among the three compounds because of its high Sd value at the low level
of cell damage. Sd could be used as a criterion for estimating the
safety of enhancers.
-16-
Primary Structure of a Squid Acid and Base Non-specific
Ribonuclease
Ayumu KUSANO, Masanori IWAMA, Kazuko OHGI, and Masachika IRIE
Ohashi Hospital, Toho University, Ohashi 20-17-6, Meguro, Tokyo 153,
Japan
Department of Microbiology, Hoshi College of Pharmacy, Ebara 2-4-41, Shinagawa,
Tokyo 142, Japan
Received July 8, 1997
Squid (Todarodes pacificus) liver RNase (RNase Tp) was purified. RNase
Tp was a base non-specific and acid RNase. Upon hydrolysis of RNA, RNase
Tp released four mononucleotides in the order of G`A`U`C. RNase Tp consisted
of two peptides with 198 and 23 amino acid residues. The amino acid sequences
of these peptides were analyzed. The large peptide had two unique segments
containing most of the active site amino acid residues of RNase T2 family
enzymes. From the comparison of the sequence of short peptide with the
sequences of the other RNase belonging to RNase T2 family RNases, it was
found that the amino acid sequence of the short peptide was very similar
to that of the C-terminal portion of RNases of the RNase T2 family. Thus,
we concluded that the short peptide was a C-terminal part of RNase Tp.
The molecular mass of the protein moiety of RNase Tp was 25,582 daltons.
The amino acid sequence of RNase Tp most resembles that of oyster RNase
(91 amino acid residues identical) in the RNase T2 family RNases. However,
the N-terminal portion of RNase Tp was unusually similar to those of plant
RNases, rather than the other animal RNases.
-17-
Phenylphenalenone-type Phytoalexins from Unripe BuEngulan
Banana Fruit
Tsunashi KAMO,1,2 Nagomi KATO,1 Nobuhiro HIRAI,1,2, Mitsuya TSUDA,3 Daie FUJIOKA,4 and Hajime OHIGASHI1
1Division of Applied Life Sciences, Graduate School of Agriculture,
Kyoto University, Kitashirakawa-oiwake-cho, Sakyo-ku, Kyoto 606-01, Japan
2CREST, Japan Science and Technology Corporation (JST)
3Division of Environmental Science and Technology, Graduate School
of Agriculture, Kyoto University, Kitashirakawa-oiwake-cho, Sakyo-ku, Kyoto
606-01, Japan
4Osaka Co-operative Wholesale Society, 3-3-3 Tenjinbashi, Kita-ku,
Osaka 530, Japan
Received July 8, 1997
Fourteen phenylphenalenone-type phytoalexins (1-14), including three new
compounds, were isolated from the peel of unripe Musa acuminata [AAA] cv.
BuEngulan fruit which had been injured and then inoculated with conidia
of Colletotrichum musae. These new phytoalexins were identified as ({)-cis-2,3-dihydro-2,3-dihydroxy-4-(4?-hydroxyphenyl)phenalen-1-one
(12), 9-(3?,4?-dimethoxyphenyl)-2-methoxyphenalen-1-one (13) and 9-(4?-hydroxyphenyl)-2-methoxyphenalen-1-one
(14). The ratios of the relative intensities of the [M]{E[M-H]{
ions or [M-H2O]{E[M-H2O-H]{ ions in the EI mass spectra
were applied to discriminate between 4- and 9-phenylphenalenones. An antifungal
test on the phytoalexins showed that a phenolic hydroxyl group was essential
for the activity.
-18-
Near Infra Red Detection of Internally Moldy Nuts
Susumu HIRANO, Noriyuki OKAWARA, and Shuichirou NARAZAKI
Quality Assurance Division, Morinaga and Co., Ltd., Shimosueyoshi 2-1-1, Tsurumiku, Yokohama 230, Japan
Received July 14, 1997
Transmittance near infra red (NIR) spectra (500-1500 nm) of individual
peanut was measured to detect the internally moldy nuts. The moldy nuts
the appearance of which had little difference from the sound nuts by visual
observations could be distinguished from each other by comparing the transmittance
ratio of 700 nm to 1100 nm by NIR spectrometry. The fungal hydrolysis of
the triglycerides that were contained in the nut seemed to account for
these differences on the NIR spectra. Because of the higher incidence of
aflatoxin (AF) contamination on these moldy nuts, taking out the internally
moldy nuts detected by NIR, could drastically reduce the AF content of
the overall lot of peanuts.
-19-
Potassium Iodide-Induced Changes in Pyruvate Dehydrogenase
Complex from Bacillus stearothermophilus
Yoichi ASO, Yasuaki HIROMASA, Yoshikatsu AIKAWA, Kohji MENO, and Masatsune ISHIGURO
Laboratory of Protein Chemistry and Engineering, Graduate School of Genetic Resources Technology, Kyushu University, Fukuoka 812-81, Japan
Biophysics Laboratory, The Institute of Physical and Chemical Research (RIKEN), Wako, Saitama 351-01, Japan
Recieved July 15, 1997
Treatment with KI and its subsequent removal induced disassembly of Bacillus
stearothermophilus pyruvate dehydrogenase complex (PDC) and association
of disassembly products, respectively. The disassembly yielded neither
completely dissociated components nor aggregate, but did yield a few molecular
forms smaller than PDC. Depending on the KI concentration, these changes
were of three phases: K-1, below 0.6 M; K-2, 1.0-1.5 M; K-3, above 1.8
M. PDC was disassembled in K-1 to C1 comprising pyruvate decarboxylase
and lipoate acetyltransferase mainly and C2 comprising the decarboxylase
and dihydrolipoamide dehydrogenase. In K-1, the removal of KI resulted
in an apparent reconstitution of PDC. The mixing of C1 with an excess of
C2 yielded an assembly larger than PDC and restored enzyme activities,
but specific activities were different from those of PDC. In K-2 and K-3
phases, complexes smaller than PDC were yielded from disassembly products,
and activities except for that of the acetyltransferase were not restored.
-20-
Purification and Characterization of an Isomaltotriose-producing
Endo-dextranase from a Fusarium sp.
Eishun SHIMIZU, Takehiro UNNO,* Manabu OHBA,** and Gentaro OKADA**,
Novo Nordisk Bioindustry Ltd., Chiba 261-01, Japan
*Research Institute, Nihon Shokuhin Kako Co., Ltd., Fuji 417, Japan
**Department of Biology, Faculty of Education, Shizuoka University, Shizuoka
422, Japan
Received July 18, 1997
An isomaltotriose-producing endo-dextranase was simply purified from cell-free
culture broth of a Fusarium sp. by ethanol fractionation and consecutive
column chromatographies using DEAE-Toyopearl and Bio-Gel P-100. The purified
enzyme was judged to be homogeneous on PAGE and SDS-PAGE as well as isoelectric
focusing. The molecular mass of the enzyme was estimated to be about 69
kDa by SDS-PAGE. The enzyme is an acidic protein with a pI of 4.6. The
optimum pH and temperature were pH 6.5 and 35EC, respectively. The enzyme
was completely stable over the range of pH 4.5-11.8 at 4EC for 24 h and
at temperatures below 45EC. Inactivation of the enzyme was found to be
partial with 5 mM Cu2{, being about 70% inhibition and complete with
5 mM of Fe3{, Hg2{, Ag{ or NBS.
@The enzyme split dextran in an endo-lytic action to produce a large amount
of isomaltotriose and a slight amount of isomaltose and glucose. The anomeric
configurations of the reaction products formed by the enzyme were a-form,
indicating that the a-glycoside linkages in the substrate are retained.
The final yield of isomaltotriose from dextran T-2000 was about 62%.
-21-
Glucan Binding Regions of Dextransucrase from Leuconostoc
mesenteroides NRRL B-512F
Kazumi FUNANE, Tetsuya OOKURA, and Mikihiko KOBAYASHI
National Food Research Institute, 2-1-2 Kannondai, Tsukuba, Ibaraki 305, Japan
Received July 18, 1997
We isolated glucan-binding peptides of a dextransucrase from Leuconostoc
mesenteroides B-512F. The dextransucrase was bound to DEAE-Sephadex A-50,
Sephadex G-100, and mutan from Streptococcus mutans. Mild trypsin digestion
dissociated the enzyme and glucan binding. In the presence of ammonium
sulfate, several peptides were bound to glucan after trypsin digestion.
Four main mutan-binding peptides were obtained by this method, and those
amino acid sequences were analyzed. One of them was identical with the
dextran-binding peptide that contains lysine, which was previously isolated
by differential chemical modification with o-phthalaldehyde. We also found
mutan-binding peptides in sucrose- and dextran-binding regions and a lysine-rich
region. Also, there was a peptide similar in sequence to glucan-binding
A-repeat of streptococcal glucosyltransferases.
-22-
Chitin Binding Protein (CBP21) in the Culture Supernatant
of Serratia marcescens 2170
Kazushi SUZUKI,1 Megumi SUZUKI,1 Mayumi TAIYOJI,1 Naoki NIKAIDOU,1,2 and Takeshi WATANABE1,2
1Department of Biosystem Science, Graduate School of Science and
Technology, Niigata University, Ikarashi, Niigata 950-21, Japan
2Department of Applied Biological Chemistry, Faculty of Agriculture,
Niigata University, Ikarashi, Niigata 950-21, Japan
Received July 28, 1997
A chitin binding protein (CBP21) 21 kDa in size, is a major protein in
the culture supernatant when Serratia marcescens 2170 is grown in the presence
of chitin. The gene (cbp) for CBP21 was found to be located in a region
1.5 kb downstream of the chiB gene encoding chitinase B. The cbp gene encodes
a polypeptide of 197 amino acids with a calculated size of 21.6 kDa containing
a putative signal sequence of 27 amino acids. Comparison of the amino acid
sequence of the deduced polypeptide with that of other proteins showed
that CBP21 is similar (45.3% amino acid identity) to CHB1 of Streptomyces
olivaceoviridis. Purified CBP21 prepared from the periplasmic fraction
of Esch-erichia coli carrying the cloned cbp gene showed itshighest binding
activity to squid chitin (-chitin) followed by colloidal chitin and regenerated
chitin. Binding of CBP21 to regenerated chitin was affected by pH, in particular,
low pH reduced binding activity markedly.
@The presence of similar chitin binding proteins in the distantly related
microorganisms, Streptomyces and Serratia, suggests a wide distribution
of this type of chitin binding protein in chitinolytic microorganisms.
-23
Expression of Biologically Active Human Calpastatin in
Baculovirus-infected Insect Cells and in Escherichia coli
Kiyotaka HITOMI, Akiko YOKOYAMA, and Masatoshi MAKI
School of Agricultural Sciences, Nagoya University, Chikusa, Nagoya 464-01, Japan
Recieved July 28, 1997
Calpastatin, an endogeneous inhibitor protein acting on calpain (Ca2{-dependent
cysteine proteinase), is widely distributed in animal tissues and cells.
Two different expression systems, baculovirus-infected Spodoptera frugiperda
(Sf9) insect cells and Escherichia coli, were used for overexpression of
the human calpastatin tagged with N-terminal hexahistidine peptide. Recombinant
calpastatin was purified to homogeneity by nickel ion affinity chromatography
and gel filtration separation. Purified recombinant proteins from both
systems have similar inhibitory activity for calpain.
-24-
Note
Effects of Bay m 1099, an a-Glucosidase Inhibitor, on Starch Degradation
in Germinating Mung Beans
Yotaro KONISHI, Michio AITANI, and Nobuji NAKATANI
Department of Food and Nutrition, Fuculty of Human Life Science, Osaka City University, 3-3-138, Sugimoto, Sumiyoshi-ku, Osaka 558, Japan
Received May 30, 1997
To examine the mechanism of starch degradation in legume cotyledons and
the physiological role of a-glucosidase, mung bean seeds were germinated
in the presence of Bay m 1099, an a-glucosidase inhibitor. Bay m 1099 (10
gEml medium), which minimized the growth deterioration of the mung bean
seedlings, caused no changes in the overall rate of starch degradation
and of soluble carbohydrate production in the cotyledons, although a-glucosidase
activity had been completely suppressed. Total amylase and phosphorylase
activities were not influenced by Bay m 1099. These results suggest that
the mung bean a-glucosidase is less responsible for starch degradation,
unlike wheat a-glucosidase [Konishi et al., Biosci. Biotech. Biochem.,
58, 135-139 (1994)].
-25-
Silk Protein, Sericin, Inhibits Lipid Peroxidation and
Tyrosinase Activity1
Norihisa KATO,*,2 Seiji SATO,* Atsushi YAMANAKA,* Hideyuki YAMADA,** Naozumi FUWA,** and Masakazu NOMURA**
*Department of Applied Biochemistry, Faculty of Applied Biological Science,
Hiroshima University, Higashi-Hiroshima 739, Japan
**Seiren Co. Ltd., 10-1, Keya 1-Chome, Fukui 918, Japan
Received June 2, 1997
This study provided the first evidence for an antioxidant action of the
silk protein sericin by showing that sericin suppressed in vitro lipid
peroxidation. Furthermore, sericin was found to inhibit tyrosinase activity.
These results suggest that sericin may be a valuable natural ingredient
for food and cosmetic industries.
-26-
Note
Molecular Cloning and Characterization of a cDNA Encoding Asparagine Synthetase
from Soybean (Glycine max L.) Cell Cultures
Hiroshi YAMAGATA,*, Ayumi NAKAJIMA,* Chris BOWLER,** and Teruo IWASAKI*
*Laboratory of Biochemistry, Faculty of Agriculture, Kobe University,
Nada, Kobe 657, Japan
**Stazione Zoologica, Villa Comunale, I 80121 Naples, Italy
Recieved June 11, 1997
A cDNA encoding glutamine-dependent asparagine synthetase was isolated
from dark-adapted Glycine max cell culture. The deduced amino acid sequence
showed 76-89% identity with other plant sequences. The gene for asparagine
synthetase is expressed predominantly in shoots as compared to roots of
etiolated plants and the level of expression decreases following light
treatment, suggesting that the gene expression is down-regulated by light.
-27-
Note
Increase of Foreign Gene Expression in Monocot and Dicot Cells by an Intron
in the 5? Untranslated Region of a Soybean Phosphoenolpyruvate Carboxylase
Gene
Tomohiko KATO, Emiko ITOH, Robert F. WHITTIER, and Daisuke SHIBATA
Mitsui Plant Biotechnology Research Institute, TCI-A1, Sengen 2-1-6, Tsukuba, Ibaraki 305, Japan
Received June 13, 1997
A genomic clone containing part of the coding region and upstream sequences
of a phosphoenolpyruvate carboxylase (PEPC) gene was isolated from a soybean
genomic library. The first intron of this gene is located in the 5? untranslated
region. This intron-spanning fragment is capable of increasing GUS activity
in plant cells.
-28-
Note
Purification and Characterization of the 2-Ketoaldonate Reductase from
Brevibacterirnm ketosoreductum ATCC21914
Do-Young YUM, Sung-Sook BAE, and Jae-Gu PAN
Bioprocess Engineering Division, Korea Research Institute of Bioscience and Biotechnology (KRIBB), P. O. Box 115, Yusong, Taejon 305-600, Korea
Received June 13, 1997
2-Ketoaldonate reductase, which is involved in ketogluconate catabolism,
was purified to homogeneity from Brevibacterium ketosoreductum ATCC21914.
The enzyme was found to catalyze the reduction of 2,5-diketo-D-gluconate
to 5-keto-D-gluconate, and to a lesser extent, 2-keto-D-gluconate to D-gluconate,
and 2-keto-L-gulonate to L-idonate. The molecular mass of the reductase
was 35 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis
and 72 kDa by gel filtration, indicating that the native enzyme may exist
as a dimer. The reductase was optimally active at pH 6.0 with NADPH as
a preferred electron donor. The pI of 4.7 was measured for the enzyme.
The apparent Km for 2,5-diketo-D-gluconate and NADPH were 5 M and 10
M, respectively. The amino-terminal amino acid sequence was NH2-Ala-Ser-Ile-Ser-Val-Ser-Val-Pro-Ser-Ala-Arg-Leu-Ala-Glu-Asp-Leu-Ser-Asp-Ile-Glu.
-29-
Note
Effect of Antioxidants in Preventing the Thermal Decomposition of Phosphatidylcholine
Hydroperoxide
Naomichi BABA, Ikue TAKAHASHI, Hiroko DAIDO, Md. Khorshed ALAM, Shuhei NAKAJIMA, and Takao KANEKO*
Department of Bioresources Chemistry, Faculty of Agriculture, Okayama
University, Tsushimanaka, Okayama 700, Japan
*Tokyo Metropolitan Institute of Gerontology, 35-2 Sakaecho, Itabashi-ku,
Tokyo 173, Japan
Received June 16, 1997
Some typical antioxidants, sesamol, -tocopherol, BHT and ascorbic acid,
were examined for their effects on preventing the thermal decomposition
of phosphatidylcholine hydroperoxide (PC-OOH), sesamol alone showing some
activity. The others, however, had no significant effect, which is in sharp
contrast to our previous studies on their remarkable activity toward the
thermal decomposition of fatty acid hydroperoxide. Ascorbic acid, which
accelerated the decomposition of fatty acid hydroperoxide, was found to
have no effect on PC-OOH in its decomposition and prevention.
-30-
Note
Analysis of Glycerophospholipid Hydroperoxides by Ion Spray Mass Spectrometry
Naomichi BABA, Hiroko DAIDO, Tomoko KOSUGI, Miwako MIYAKE, and Shuhei NAKAJIMA
Faculty of Agriculture, Okayama University, 1-1-1 Tsushimanaka, Okayama 700, Japan
Received June 16, 1997
Synthetic phosphatidylcholine hydroperoxide (PC-OOH), phosphatidylethanolamine
hydroperoxide (PE-OOH) and phosphatidylglycerol hydroperoxide (PG-OOH)
could be analyzed by ion spray ionization mass spectrometry, clearly affording
the molecular ion peaks as protonated ion species without any fragmentation.
Linearity between PC-OOH concentration and peak intensity, and the minimum
detectable concentration of PC-OOH were obtained.
-31-
Note
Effects of Unsaturated Uronic Acid Residues at Non-reducing End on Bond
Cleavage Frequency of Poly(1,4--L-guluronide) Lyase from Enterobacter
cloacae M-1
Tomoko SHIMOKAWA, Shigeki YOSHIDA, and Isao KUSAKABE
Institute of Applied Biochemistry, University of Tsukuba, 1-1-1 Tennoodai, Tsukuba-shi, Ibaraki 305, Japan
Received June 20, 1997
The mode of action of poly(1,4-a-L-guluronide) lyase from Enterobacter
cloacae M-1 on unsaturated oligoguluronic acids was studied using fluorophore-assisted
carbohydrate electrophoresis. The polyguluronate lyase degraded unsaturated
penta-, hexa-, and heptaguluronic acids, but not unsaturated oligoguluronic
acids with DPs less than 4. On comparison with the aspect of enzymatic
degradation of unsaturated oligoguluronic acid and saturated oligoguluronic
acid having the same DP, the former was degraded faster than the latter,
and also the cleavage pattern of the polyguluronate lyase on unsaturated
oligoguluronic acids was considerably different from that on saturated
oligoguluronic acids. From the results described above, we suggest that
the affinity of the first subsite from the non-reducing end side of the
enzyme to residues is lower than that to GulA residues.
-32-
Note
Escherichia coli Transformant Expressing the Glucose Dehydrogenase Gene
from Bacillus megaterium as a Cofactor Regenerator in a Chiral Alcohol
Production System
Michihiko KATAOKA, Luh Poni Sri ROHANI, Masaru WADA, Keiko KITA,* Hideshi YANASE,* Itaru URABE,** and Sakayu SHIMIZU
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto
University, Sakyo-ku, Kyoto 606-01, Japan
*Department of Biotechnology, Faculty of Engineering, Tottori University,
4-101 Koyama, Tottori 680, Japan
**Department of Biotechnology, Faculty of Engineering, Osaka University,
2-1 Yamadaoka, Suita, Osaka 565, Japan
Received July 2, 1997
Escherichia coli JM109 (pGDA2) overexpressing the glucose dehydrogenase
(GDH) gene from Bacillus megaterium IWG3 was examined for use as a cofactor
regenerator. In the asymmetric reduction of ethyl 4-chloro-3-oxobutanoate
by E. coli JM109 (pKAR) which is an aldehyde reductase-overproducing transformant,
E. coli JM109 (pGDA2) can act as an NADPH regenerator with NADP{ and
glucose, similarly to commercially available GDH.
-33-
Note
Effects of Substrate Solubility in Interesterification with Triolein by
Immobilized Lipase in Supercritical Carbon Dioxide
Seung-Heon YOON,1,2 Hideki NAKAYA,1 Osamu ITO,1 Osato MIYAWAKI,1 Kwan-Hwa PARK,2 and Kozo NAKAMURA1
1Department of Applied Biological Chemistry, Division of Agriculture
and Agricultural Life Science, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku,
Tokyo 113, Japan
2Department of Food Science and Technology, College of Agriculture
and Life Sciences, Seoul National University, 134 Seodun-dong, Kwonsun-ku,
Suwon 441-744, Korea
Received July 7, 1997
The interesterification reaction by immobilized lipase between triolein
and behenic acid (BA) or ethyl behenate (EB) was investigated in supercritical
carbon dioxide (SCCO2) to produce 1,3-dibehenoyl-2-oleoyl glycerol
(BOB). The incorporation rate of behenoyl group to triolein was much higher
with EB as a substrate than with BA. The solubility in SCCO2 was found
to be significantly higher for EB than BA, which seemed to cause the higher
formation rate of the behenoyl-enzyme complex in the former case to give
the higher production rate of the final interesterification product, BOB.
-34-
Note
Pyricuol, a New Phytotoxin from Magnaporthe grisea
Jin-Cheol KIM, Ji-Young MIN, Heung-Tae KIM, Kwang-Yun CHO, and Seung-Hun YU*
Screening Division, Korea Research Institute of Chemical Technology,
100 Jang-Dong, Yusong-Gu, Taejon 305-600, Korea
*Department of Agricultural Biology, College of Agriculture, Chungnam National
University, 220 Gung-Dong, Yusong-Gu, Taejon 305-764, Korea
Received July 14, 1997
A new pyriculol-related phytotoxin, designated as pyricuol (1), was isolated
from a liquid culture of Magnaporthe grisea, the causal fungus of rice
blast disease, together with two known metabolites, pyriculol (2) and dihydropyriculol.
Its structure was determined on the basis of physicochemical and spectroscopic
data to be 2-(3-hydroxymethyl-1,4-hexadienyl)-6-hydroxybenzaldehyde.
-35-
Note
Evaluation of the Antioxidative Activity of Tea by an Oxygen Electrode
Method
Midori KUMAMOTO and Tamiyoshi SONDA
Department of Food and Nutrition, Faculty of Domestic Science, Nishikyushu University, 4490-9 Kanzaki-machi, Kanzaki-gun, Saga 842, Japan
Received July 17, 1997
The antioxidative activity was studied for 25 kinds of tea and catechins
by a new evaluation method using an oxygen electrode. The concentration
of catechins in 6 types of green tea was analyzed by HPLC. The result indicates
that the antioxidative activity of green tea depends to some extent on
the amount of catechins present.
-36-
Note
Inhibition of Glucan Synthesis by Casein Polymers Crosslinked by Glutaraldehyde
Osamu HAYASHIDA, Fumiki MORI, Keiji HASUMI, and Akira ENDO
Department of Applied Biological Science, Faculty of Agriculture, Tokyo Noko University, Fuchu-Shi, Tokyo 183, Japan
Received July 17, 1997
Glutaraldehyde-crosslinked a-casein polymers with molecular masses of 1-5~105
were found to inhibit adhesive insoluble glucan formation catalyzed by
glucan synthases from Streptococcus sobrinus B13, a cariogenic oral bacterium.
Of the three subcomponents of the glucan synthases tested, primer-dependent
insoluble glucan synthase was inhibited specifically by the MrE2.7~105
polymer at a concentration of 1.4-8.5 gEml. The polymer was also active
in inhibiting the artificial plaque formation by S. sobrinus B13.
-37-
Note
Reduction of Alkyl (2-Oxocyclohexyl)acetates by Baker's Yeast
Makoto GANAHA, Yuhei FUNABIKI, Minoru MOTOKI, Satoshi YAMAUCHI, and Yoshiro KINOSHITA
Department of Agricultural Chemistry, Faculty of Agriculture, Ehime University, Matsuyama 790, Japan
Received July 28, 1997
Baker's yeast reduction of methyl and ethyl (2-oxocyclohexyl)acetates proceeded
with enantio- and diastereo-selectivity, affording the corresponding (2S)-trans-alcohols
(major), (2S)-cis-alcohols (minor), and the unaltered (1S)-ketones with
high optical purity.
-38-
Note
The Characterization of Acetic Acid Bacteria Efficiently Producing Bacterial
Cellulose from Sucrose: The Proposal of Acetobacter xylinum subsp. nonacetoxidans
subsp. Nov.
Yukiko KOJIMA, Naoto TONOUCHI, Takayasu TSUCHIDA, Fumihiro YOSHINAGA, and Yuzo YAMADA
Bio-Polymer Research Co., Ltd., Takatsu-ku, Kawasaki 213, Japan
*Laboratory of Applied Microbiology, Department of Applied Biological Chemistry,
Faculty of Agriculture, Shizuoka University, Shizuoka 422, Japan
Received July 28, 1997
A taxonomic study was done for the isolates obtained as cellulose high
producers from sucrose. These strains were found to have a common characteristic
unique for Acetobacter; they did not oxidize acetate and lactate. Therefore,
we concluded that these isolates are classified as a new subspecies and
proposed Acetobacter xylinum subsp. nonacetoxidans subsp. nov. BPR 2002
(JCM 10150) was designated as a type strain of the new subspecies.
-39-
Note
Simple Method for Detecting Glycoproteins Dot-blotted or Electro-blotted
on to a Polyvinylidene Difluoride Membrane
Shizuya KABUTO and Koji MURAMOTO
Department of Biological Resource Sciences, Graduate School of Agriculture, Tohoku University, Sendai 981, Japan
Received July 30, 1997
A simple method is described for detecting glycoproteins which had been
dot-blotted or electro-blotted on to a polyvinylidene difluoride (PVDF)
membrane after sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis.
This method is based on the periodate oxidation of glycoproteins on a PVDF
membrane with subsequent staining by the chromophoric hydrazide, 4-N,N-dimethylamino-4?-azobenzene
sulfonyl hydrazide. The glycoproteins could be visualized as red-colored
spots or bands in the range of 0.16-0.31 g.
-40-
Note
Novel Stereoselective Reaction of Levoglucosenone with Furfural
Toshio NISHIKAWA, Hiroshi ARAKI, and Minoru ISOBE*
Laboratory of Organic Chemistry, School of Bioagricultural Sciences, Nagoya University, Chikusa, Nagoya 464-01, Japan
Received August 13, 1997
Levoglucosenone reacted with furfural in the presence of an aqueous base
to give a product in high yield with high stereoselectivity. The structure,
including the stereochemistry of the product, was elucidated by NMR analyses.
-41-
Short Communication
Compensation for D-Glutamate Auxotrophy of Escherichia coli WM335 by D-Amino
Acid Aminotransferase Gene and Regulation of murI Expression
Lidong LIU,1 Tohru YOSHIMURA,1 Keiji ENDO,1 Kazuhisa KISHIMOTO,1 Yoshihiro FUCHIKAMI,1 James M. MANNING,2 Nobuyoshi ESAKI,1 and Kenji SODA3
1Institute for Chemical Research, Kyoto University, Gokasho, Uji,
Kyoto-fu 611, Japan
2Department of Biology, Northeastern University, 414 Mugar Building,
Boston, MA, USA
3Faculty of Engineering, Kansai University, Yamate-cho, Suita, Osaka-fu
564, Japan.
Received June 13, 1997
D-Glutamate, an indispensable component of peptidoglycans of bacteria,
is provided by glutamate racemase in E. coli cells. Compensation for D-glutamate
auxotrophy of E. coli WM335 cells lacking the glutamate racemase gene,
murI, with the D-amino acid aminotransferase gene suggests the presence
of a threshold concentration for the D-glutamate required by E. coli cells,
as well as a regulation system for murI expression.