(Vol.68 No.5 2004)
Review
Bacterial Two-component and Hetero-heptameric Pore-forming Cytolytic Toxins:
Structures, Pore-forming Mechanism, and Organization of the Genes
Jun Kaneko and Yoshiyuki Kamio† p.981
A New Assay Using Surface Plasmon Resonance (SPR) to Determine
Binding of the Lactobacillus acidophilus Group to Human Colonic Mucin
Hideaki Uchida,1 Kenji Fujitani,1 Yasushi Kawai,1 Haruki Kitazawa,1 Akira Horii,2
Kenichi Shiiba,3 Kazuya Saito,3 and Tadao Saito1,† p.1004
Effect of Difructose Anhydride III on Calcium Absorption in Humans
Norihiro Shigematsu,1,† Yasuhide Okuhara,1 Takuya Shiomi,1 Fusao Tomita,2,*
and Hiroshi Hara2 p.1011
Elevation of Intracellular cAMP Levels by Dominant Active Heterotrimeric
G Protein Alpha Subunits ScGP-A and ScGP-C in Homobasidiomycete, Schizophyllum
commune*
Kenji Yamagishi,1,† Toshiyuki Kimura,1 Masahiro Suzuki,1 Hiroshi Shinmoto,2
and Ko-ji Yamaki1 p.1017
Anti-microbial Action against Verotoxigenic Escherichia coli O157:H7
of Nitric Oxide Derived from Sodium Nitrite
Hidetoshi Morita,1,† Hiroshi Yoshikawa,1 Takehito Suzuki,1 Shin Hisamatsu,2
Yukio Kato,1 Ryoichi Sakata,1 Yukiharu Nagata,1 and Tetsuhiko Yoshimura3 p.1027
Effect of a Taurine Treatment on the Regression of Existing Atherosclerotic
Lesions in Rabbits Fed on a High-cholesterol Diet
Jale Balkan,1,† Serdar Öztezcan,1 Aydan Hatipoglu,2 Uğur Çevikbaş,3
Gülçin Aykaç-toker,1 and Müjdat Uysal1
p.1035
Evaluation of the Preventive Effect of Isoflavone Extract on Bone
Loss in Ovariectomized Rats
Yoon-Bok Lee,1,2 Hyong Joo Lee,2,† Kang Sung Kim,3 Jae-Yong Lee,4 Sang-Yoon
Nam,5 Sang-Hee Cheon,1 and Heon-Soo Sohn1 p.1040
Improving the Pyrophosphate-inosine Phosphotransferase Activity
of Escherichia blattae Acid Phosphatase by Sequential Site-directed Mutagenesis
Yasuhiro Mihara,1,† Kohki Ishikawa,2 Ei-ichiro Suzuki,2 and Yasuhisa Asano3
p.1046
Isoflavones Found in Korean Soybean Paste as 3-Hydroxy-3-methylglutaryl
Coenzyme A Reductase Inhibitors
Jang Hoon Sung,1 Seung Jun Choi,1 Soon Won Lee,2 Kwan Hwa Park,1 and Tae Wha
Moon1,† p.1051
Characterization of Endo-β-N-acetylglucosaminidase from Alkaliphilic
Bacillus halodurans C-125
Kiyotaka Fujita,1 Hideto Takami,2 Kenji Yamamoto,1 and Kaoru Takegawa3,†
p.1059
Dietary Branched-chain Amino Acids Suppress the Expression of
Pancreatic Amylase mRNA in Rats
Naoto Hashimoto and Hiroshi Hara† p.1067
Enzymatic Properties and Nucleotide and Amino Acid Sequences of
a Thermostable β-Agarase from the Novel Marine Isolate, JAMB-A94
Yukari Ohta, Yuichi Nogi, Masayuki Miyazaki, Zhijun Li, Yuji Hatada, Susumu
Ito,† and Koki Horikoshi p.1073
Purification, Cloning, and Sequence Analysis of β-N-Acetylglucosaminidase
from the Chitinolytic Bacterium Aeromonas hydrophila Strain SUWA-9
Xiqian Lan, Naomi Ozawa, Naohide Nishiwaki, Ritsuko Kodaira, Mitsuo Okazaki,
and Makoto Shimosaka† p.1082
Gelling Properties of Soybean β-Conglycinin Having Different
Subunit Compositions
Mohamad ramlan Bin Mohamed Salleh,1,2 Nobuyuki Maruyama,1 Koji Takahashi,3 Kazuhiro
Yagasaki,4 Takahiko Higasa,1 Yasuki Matsumura,1 and Shigeru Utsumi1,†
p.1091
Synthesis of Brassinosteroids of Varying Acyl Side Chains and
Evaluation of Their Brassinolide-like Activity
Shinya Uesusuki, Bunta Watanabe, Shuji Yamamoto, Junko Otsuki, Yoshiaki Nakagawa,†
and Hisashi Miyagawa p.1097
Isolation of Flavohemoglobin from the Actinomycete Streptomyces
antibioticus Grown without External Nitric Oxide Stress
Yasuyuki Sasaki,1 Naoki Takaya,1 Akira Nakamura,1 and Hirofumi Shoun2,†
p.1106
Effects of Truncation at the Non-homologous Region of a Family
3 β-Glucosidase from Agrobacterium tumefaciens
Li Ying,1,2 Motomitsu Kitaoka,1,† and Kiyoshi Hayashi1 p.1113
Identification of High Molecular Weight Proteins in Squid Muscle
by Western Blotting Analysis and Postmortem Rheological Changes
Chinatsu Kasamatsu,1,2,† Sumiko Kimura,3 Mieko Kagawa,4 and Keiko Hatae1
p.1119
Novel Fusicoccins R and S, and the Fusicoccin S Aglycon (Phomopsiol)
from Phomopsis amygdali Niigata 2-A, and Their Seed Germination-stimulating
Activity in the Presence of Abscisic Acid
Naoto Tajima,1 Manabu Nukina,1,2 Nobuo Kato,3 and Takeshi Sassa1,2,† p.1125
Note
New Monoterpene Glucoside from the Aerial Parts of Thyme (Thymus vulgaris L.)
Hiroyuki Takeuchi,1,† Zhan-Guo Lu,1 and Tomoyuki Fujita2 p.1131
Note
Suppressive Effects of Dietary Fiber in Yogurt on the Postprandial Serum Lipid
Levels in Healthy Adult Male Volunteers
Shizuki Kondo, Jin-zhong Xiao,† Noritoshi Takahashi, Kazuhiro Miyaji,
Keiji Iwatsuki, and Sadayuki Kokubo p.1135
Note
Design of Soymetide-4 Derivatives to Potentiate the Anti-alopecia Effect
Takahiro Tsuruki and Masaaki Yoshikawa† p.1139
Note
Enzymatic Reactions by Five Chalcone Synthase Homologs from Hop (Humulus lupulus
L.)
Yukio Okada,1,† Yukie Sano,2 Takafumi Kaneko,1 Ikuro Abe,2 Hiroshi Noguchi,2
and Kazutoshi Ito1 p.1142
Note
Isolation of an Exopolysaccharide-producing Bacterium, Sphingomonas sp. CS101,
Which Forms an Unusual Type of Sphingan
Eun-Jung Seo,1,* Sang-Ho Yoo,2,* Ko-Woon Oh,1 Jaeho Cha,3 Hyeon Gyu Lee,4 and
Cheon-Seok Park1,† p.1146
Note
Catalytic Activity of Tripeptidase from Lactococcus lactis to Which Amino Acid
Substitution Was Introduced According to Natural Mutation
Sumiko Mori, Satoshi Kaneko, and Takafumi Kasumi† p.1149
Note
Effects of Soybean β -Conglycinin on Hepatic Lipid Metabolism and Fecal
Lipid Excretion in Normal Adult Rats
Kensuke Fukui,1,† Makiko Kojima,1 Nobuhiko Tachibana,1 Mitsutaka Kohno,1
Kiyoharu Takamatsu,1 Motohiko Hirotsuka,1 and Makoto Kito2 p.1153
Note
Involvement of a Glu71–Arg64 Couple in the Access Channel for NADH in
Cytochrome P450nor
Fei Su,1 Shinya Fushinobu,2 Naoki Takaya,1 and Hirofumi Shoun2,† p.1156
Note
Resistance Imparted by Traditional Chinese Medicines to the Acute Change of
Glutamic Pyruvic Transaminase, Alkaline Phosphatase and Creatine Kinase Activities
in Rat Blood Caused by Noise
Bei-Wei Zhu,1 Yu-Mei Sun,1 Xia Yun,1 Song Han,2 Mei-Lan Piao,2 Yoshiyuki Murata,3
and Mikiro Tada2,† p.1160
Note
Biotransformation of α-Isomethylionone to 1-(2,6,6-Trimethyl-2-cyclohexen-1-yl)propan-2-one
Susumu Ishizaki,1 Masamichi Itoh,1,† Tsuyoshi Komai,1 Tsutomu Honda,1
and Takeshi Kitahara2 p.1164
Note
Microbial Hydroxylation of Indole to 7-Hydroxyindole by Acinetobacter calcoaceticus
Strain 4-1-5
Daisuke Sugimori,1,† Takanori Sekiguchi,1 Fumihiko Hasumi,2 Motoki Kubo,3
Naoki Shirasaka,4 and Masaya Ikunaka4 p.1167
Communication
Characterization of Protein Phosphatase 2A Acting on Phosphorylated Plasma Membrane
Aquaporin of Tulip Petals
Abul Kalam Azad,1,2 Yoshihiro Sawa,1 Takahiro Ishikawa,1 and Hitoshi Shibata1,†
p.1170
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Review
Bacterial Two-component and Hetero-heptameric Pore-forming Cytolytic Toxins:
Structures, Pore-forming Mechanism, and Organization of the Genes
Jun Kaneko and Yoshiyuki Kamio†
Laboratory of Applied Microbiology, Department of Microbial Biotechnology, Division of Bioscience and Biotechnology for Future Bioindustry, Graduate School of Agricultural Science, Tohoku University, Tsutsumi-dori Amamiya-machi 1-1, Aoba-ku, Sendai 981-8555, Japan
Staphylococcal γ-hemolysin (Hlg), leukocidin (Luk), and Panton–Valentine leukocidin (PVL) are two-component and hetero-oligomeric pore-forming cytolytic toxins (or cytolysin), that were first identified in bacteria. No information on the existence of hetero-oligomeric pore-forming cytolytic toxins in bacteria except for staphylococcal strains is available so far. Hlg (Hlg1 of 34 kDa/Hlg2 of 32 kDa) effectively lyses erythrocytes from human and other mammalian species. Luk (LukF of 34 kDa/LukS of 33 kDa) is cytolytic toward human and rabbit polymorphonuclear leukocytes and rabbit erythrocytes, and PVL (LukF-PV of 34 kDa/LukS-PV of 33 kDa) reveals cytolytic activity with a high cell specificity to leukocytes. Hlg1 is identical to LukF and that the cell specificities of the cytolysins are determined by Hlg2 and LukS. Based on the primary and 3-dimensional structures of the toxin components, Hlg, Luk, and PVL are thought to form a family of proteins. In the first chapter of this article, we describe the molecular basis of the membrane pore-forming nature of Hlg, Luk, and PVL. We also describe a requirement of the phosphorylation of LukS and LukS-PV by protein kinase for their leukocytolytic activity besides their pore formation on human leukocytes.
Recently, the assembly mechanism of the LukF and Hlg2 monomers into pore-forming hetero-oligomers of Hlg on human erythrocyte membranes has been clarified for the first time by our study using a single-molecular fluorescence imaging technique. We estimated 11 sequential equilibrium constants for the assembly pathway which includes the beginning with membrane binding of monomers, proceeds through single pore oligomerization, and culminates in the formation of clusters of the pores. In the second chapter of this article, we refer to an assembly mechanism of LukF and Hlg2 on human erythrocytes as well as the roles of the membranes of the target cells in pore formation by Hlg.
The LukF, LukS, and Hlg2 proteins are derived from the Hlg locus (hlg), and have been found in 99% of clinical isolates of Staphylococcus aureus. In contrast, LukF-PV and LukS-PV are derived from the PVL locus (pvl) which is distinct from the hlg locus, and only a small percentage of clinically isolated S. aureus strains carries pvl. Recently, we discovered pvl on the genome of lysogenic bacteriophages, φPVL, and determined the entire gene of the phage. We also demonstrated the phage conversion of S. aureus leading to the production of PVL through the discovery of a PVL-carrying temperate phage, φSLT, from a clinical isolate of S. aureus. In the third chapter of this article, we discuss genetic analyses of the Hlg, Luk, and PVL genes. We also discuss the current status of knowledge of the genetic organization of PVL-converting phages in order to achieve an understanding of their molecular evolution.
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A New Assay Using Surface Plasmon Resonance (SPR) to Determine Binding of the
Lactobacillus acidophilus Group to Human Colonic Mucin
Hideaki Uchida,1 Kenji Fujitani,1 Yasushi Kawai,1 Haruki Kitazawa,1 Akira Horii,2 Kenichi Shiiba,3 Kazuya Saito,3 and Tadao Saito1,†
1Laboratory of Animal Products Chemistry, Graduate School of Agricultural Science, Tohoku University, Tsutsumidori-Amamiyamachi 1-1, Aoba-ku, Sendai, Miyagi 981-8555, Japan
2Department of Molecular Pathology, Tohoku University School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan
3Department of Gastrointestinal and Colorectal Surgery, Tohoku University School of Medicine, 1-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8574, Japan
A new binding assay to investigate the mechanism of adhesion of lactic acid bacteria to the human intestine was established by the surface plasmon resonance technique using a biosensor BIACORE1000. Cells of 26 strains of the Lactobacillus acidophilus group as analytes were eluted onto a sensor chip on which were immobilized biotinylated A-trisaccharide polymer probes having human A-type antigen [(GalNAcα1-3(Fucα1-2)Gal)-] or human colonic mucin of blood type A (HCM-A) as ligands. In the first screening, high adhesive affinity to the A-trisaccharide BP-probe was observed in L. acidophilus OLL2769, L. crispatus JCM8778, LA205 and LA206. In the second screening, which used HCM-A, only L. acidophilus OLL2769 and L. crispatus JCM8778 were selected as adhesive strains with specific binding ability to human A-antigen. The results indicated that some strains of the L. acidophilus group could recognize and bind the sugar chain of A-antigen structure on HCM.
-3-
Effect of Difructose Anhydride III on Calcium Absorption in Humans
Norihiro Shigematsu,1,† Yasuhide Okuhara,1 Takuya Shiomi,1 Fusao Tomita,2,* and Hiroshi Hara2
1Food Research Division, Central Research Laboratory, FANCL Co., Ltd., 12-13 Kamishinano, Totsuka-ku, Yokohama-shi, Kanagawa-ken 244-0806, Japan
2Division of Applied Bioscience, Graduate School of Agriculture, Hokkaido University, Sapporo 060-8589, Japan
The effects of difructose anhydride III (DFAIII) on stimulating calcium absorption was investigated in humans. We studied changes in the time-course of characteristics urinary calcium excretion in 12 healthy men given 0.3, 1.0 or 3.0 g of DFAIII and 300 mg of calcium as calcium carbonate. In addition, urinary excretion and urine concentrations of creatinine and deoxypyridinoline were determined. Urine calcium excretion every 2 hours after the intake were higher over than that of the control subjects. The total amount of urinary calcium excretion for 10 hours was significantly greates in the subjects given 1.0 g or 3.0 g of DFAIII than that of the control subjects. However, there were no differences in the urine concentrations of creatinine and deoxypyridinoline between the subjects given DFAIII and the control subjects. These findings suggests that low dose of DFAIII had a stimulating effect on calcium absorption in humans.
-4-
Elevation of Intracellular cAMP Levels by Dominant Active Heterotrimeric G Protein
Alpha Subunits ScGP-A and ScGP-C in Homobasidiomycete, Schizophyllum commune*
Kenji Yamagishi,1,† Toshiyuki Kimura,1 Masahiro Suzuki,1 Hiroshi Shinmoto,2 and Ko-ji Yamaki1
1National Agricultural Research Center for the Tohoku Region, National Agricultural Research Organization, Arai, Fukushima 960-2156, Japan
2National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan
In many fungi, the heterotrimeric G protein alpha subunits, and/or small G protein (RAS) control intracellular cAMP levels. But it is not clear which types of G proteins modulate cAMP levels in homobasidiomycete (mushrooms). To explain the mechanism, we expressed dominant active RAS (a homolog of S. cerevisiae RAS1) in homobasidiomycete Schizophyllum commune and compared the cAMP levels in the transformed clones with those of clones expressing dominant active heterotrimeric G protein alpha subunits ScGP-A, B, and C. The results demonstrated that the dominant active ScGP-A and C elevated the intracellular cAMP levels. In contrast, the dominant active S. commune RAS gene did not affect the cAMP levels, even though colony growth and formation of fruiting bodies were apparently repressed. These data suggest that the heterotrimeric G protein alpha subunits are involved in the mechanism of cAMP regulation, and that RAS modulates another signal-transduction pathway regulating cell growth and differentiation.
-5-
Anti-microbial Action against Verotoxigenic Escherichia coli O157:H7 of Nitric
Oxide Derived from Sodium Nitrite
Hidetoshi Morita,1,† Hiroshi Yoshikawa,1 Takehito Suzuki,1 Shin Hisamatsu,2 Yukio Kato,1 Ryoichi Sakata,1 Yukiharu Nagata,1 and Tetsuhiko Yoshimura3
1School of Veterinary Medicine, Azabu University, Sagamihara 229-8501, Japan
2School of Environmental Health Science, Azabu University, Sagamihara 229-8501, Japan
3Institute for Life Support Technology, Yamagata Public Corporation for the Development of Industry, Matsuei, Yamagata 990-2473, Japan
The levels of verotoxin-1 and verotoxin-2 released by verotoxigenic Escherichia coli O157:H7 treated in vitro with sodium nitrite, sodium chloride and several antibiotics were evaluated. Of the three strains of E. coli O157:H7 used in this study, two strains produced both verotoxin-1 and verotoxin-2, and one strain produced only verotoxin-2. Treatment of E. coli O157:H7 with sodium nitrite (6,000 mg/l, minimum inhibitory concentration) did not increase the levels of verotoxin-1 and verotoxin-2 compared with a treatment by sodium chloride or antibiotics. When the electron paramagnetic resonance spectrum of sodium nitrite-treated bacterial cells was examined at 77 K to clarify the mechanism for the anti-bacterial activity of nitric oxide derived from sodium nitrite, electron paramagnetic resonance signals with g-values of 2.035 and 2.010 were observed. These were identified as being derived from iron–nitric oxide complexes. It appears that the dinitrosyl iron complexes in the E. coli O157:H7 cells were generated from the reaction of iron-sulfur proteins (enzymes) with nitric oxide formed by the reduction of sodium nitrite. The amount of ATP was decreased by the presence of sodium nitrite in the cell suspension. These findings indicate that nitric oxide derived from sodium nitrite penetrated the cells and inactivated enzymes related to the respiratory chain.
-6-
Effect of a Taurine Treatment on the Regression of Existing Atherosclerotic
Lesions in Rabbits Fed on a High-cholesterol Diet
Jale Balkan,1,† Serdar Öztezcan,1 Aydan Hatipoglu,2 Uğur Çevikbaş,3 Gülçin Aykaç-toker,1 and Müjdat Uysal1
1Department of Biochemistry, İstanbul Faculty of Medicine, İstanbul University, 34093 Çapa-İstanbul, Turkey
2Eczacibaşi Pharmaceutical Company, Lüleburgaz, Turkey
3Department of Pathology, İstanbul Faculty of Medicine, İstanbul University, Turkey
We studied whether taurine has any regressive effect on existing atherosclerotic lesions and lipid peroxidation in rabbits fed on a high-cholesterol (HC) diet. The cholesterol, triglyceride, malondialdehyde (MDA) and diene conjugate (DC) levels, as well as the aortic histopathological findings were examined in rabbits that had been fed on a cholesterol-containing diet for 8 months [0.5% cholesterol (w/w) for 3 months and subsequently 0.25% cholesterol (w/w) for 5 months], and then for a further 4 months on a normal diet with or without taurine treatment [1% (w/v) in the drinking water]. High levels of lipid and lipid peroxide induced by the HC diet were observed to decline in the plasma, liver and aorta of atherosclerotic rabbits, as well as a slight retardation in aortic atherosclerotic lesions during the regression period. Although no significant differences in the lipid and lipid peroxide levels in the plasma and aorta were found between the regressed groups with or without the taurine treatment, the extent of atherosclerotic lesions in the aorta was less in the taurine-treated regressed group than in the non-treated regressed group. However, the liver MDA and DC levels were lower in the regressed rabbits with the taurine treatment in the non-treated group. These results indicate that the taurine treatment may accelerate the regression of cholesterol-induced atherosclerotic lesions in rabbits without having any effect on the plasma and aorta lipid and lipid peroxide levels.
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Evaluation of the Preventive Effect of Isoflavone Extract on Bone Loss in Ovariectomized
Rats
Yoon-Bok Lee,1,2 Hyong Joo Lee,2,† Kang Sung Kim,3 Jae-Yong Lee,4 Sang-Yoon Nam,5 Sang-Hee Cheon,1 and Heon-Soo Sohn1
1Central Research Institute, Dr. Chung’s Food Co., Ltd., 1-25 Songjung-Dong, Chungjoo-Si, 361-782, Korea
2Department of Food Science and Technology and School of Agricultural Biotechnology, Seoul National University, San 56-1, Shillim-Dong, Seoul, 151-742, Korea
3Department of Food and Nutrition, Yong-in University, 470 Samga-Dong, Yongin-Si, 449-714, Korea
4College of Medicine, Hallym University, 1 Okchun-Dong, Chunchon-Si, 200-702, Korea
5College of Veterinary Medicine, Chungbuk National University, 48 Gaesin-Dong, Chungjoo-Si, 361-763, Korea
To examine a potential role for soybean phytoestrogens in postmenopausal bone loss, twenty-four 12-week-old Sprague-Dawley rats were divided randomly into 4 groups and given controlled diets for 16 weeks. The treatment groups were as followed: sham operated, ovariectomized (OVX) control, OVX + isoflavone extract (6.25 g/kg), and OVX + 17β-estradiol (4 mg/kg). OVX treatments reduced femoral and fourth lumbar vertebral bone density and mineral content ( p < 0.01), decreased uterine weight ( p < 0.01), accelerated body weight increases ( p < 0.05), and increased the activities ( p < 0.01) of both serum alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (TRAP). Supplementation with isoflavone prevented the losses of bone density and mineral content caused by OVX ( p < 0.01). Although both isoflavone and 17β-estradiol exhibited similar bone-sparing ability on the OVX-induced bone loss, the effect of isoflavone was not the same as that of 17β-estradiol on the serum ALP and TRAP, body weight increase, and uterine weight change. We concluded that dietary supplementation with soybean isoflavone can prevent postmenopausal bone loss via a different mechanism of estrogen in OVX rats.
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Improving the Pyrophosphate-inosine Phosphotransferase Activity of Escherichia
blattae Acid Phosphatase by Sequential Site-directed Mutagenesis
Yasuhiro Mihara,1,† Kohki Ishikawa,2 Ei-ichiro Suzuki,2 and Yasuhisa Asano3
1Applied Microbiology Laboratory, Fermentation and Biotechnology Laboratories, Ajinomoto Co., Inc., 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki-shi 210-8681, Japan
2Institute of Life Sciences, Ajinomoto Co., Inc., 1-1 Suzuki-cho, Kawasaki-ku, Kawasaki-shi 210-8681, Japan
3Biotechnology Research Center, Toyama Prefectural University, 5180 Kurokawa, Kosugi, Toyama 939-0398, Japan
Escherichia blattae acid phosphatase/phosphotransferase (EB-AP/PTase) exhibits C-5’-position selective pyrophosphate-nucleoside phosphotransferase activity in addition to its intrinsic phosphatase. Improvement of its phosphotransferase activity was investigated by sequential site-directed mutagenesis. By comparing the primary structures of higher 5’-inosinic acid (5’-IMP) productivity and lower 5’-IMP productivity acid phosphatase/phosphotransferase, candidate residues of substitution were selected. Then a total of 11 amino acid substitutions were made with sequential substitutions. As the number of substituted amino acid residues increased, the 5’-IMP productivity of the mutant enzyme increased, and the activity of the 11 mutant phosphotransferases of EB-AP/PTase reached the same level as that of Morganella morganii AP/PTase. This result shows that Leu63, Ala65, Glu66, Asn69, Ser71, Asp116, Thr135, and Glu136, whose relevance was not directly established by structural analysis alone, also plays an important role in the phosphotransferase activity of EB-AP/PTase.
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Isoflavones Found in Korean Soybean Paste as 3-Hydroxy-3-methylglutaryl Coenzyme
A Reductase Inhibitors
Jang Hoon Sung,1 Seung Jun Choi,1 Soon Won Lee,2 Kwan Hwa Park,1 and Tae Wha Moon1,†
1Department of Food Science and Technology, School of Agricultural Biotechnology and Center for Agricultural Biomaterials, Seoul National University, Seoul 151-742, Korea
2Department of Chemistry, College of Natural Science, Sungkyunkwan University, Suwon 440-746, Korea
3-Hydroxy-3-methylglutaryl CoA (HMG-CoA) reductase is the rate-limiting enzyme in the biosynthesis of cholesterol in mammals. Some microbial metabolites have been found to be HMG-CoA reductase inhibitors. Korean soybean paste is a unique food fermented by many microorganisms. The enzymatic method using the catalytic domain of Syrian hamster HMG-CoA reductase was employed for the screening of HMG-CoA reductase inhibitors. Soybean paste extract was fractionated by vacuum liquid chromatography. Fractions showing relatively high HMG-CoA reductase inhibition were further purified through Sephadex LH-20 column chromatography and C18 preparative HPLC, and the inhibitory compounds were identified as genistein, daidzein, and glycitein.
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Characterization of Endo-β-N-acetylglucosaminidase from Alkaliphilic Bacillus
halodurans C-125
Kiyotaka Fujita,1 Hideto Takami,2 Kenji Yamamoto,1 and Kaoru Takegawa3,†
1Division of Integrated Life Science, Graduate School of Biostudies, Kyoto University, Kyoto 606-8502, Japan
2Japan Marine Science and Technology Center, Microbial Genome Research Group, 2-15 Natsushima, Yokosuka, Kanagawa 237-0061, Japan
3Department of Life Sciences, Faculty of Agriculture, Kagawa University, Miki-cho, Kagawa 761-0795, Japan
The genome sequencing project on alkaliphilic Bacillus halodurans C-125 revealed a putative endo-β-N-acetylglucosaminidase (Endo-BH), which consists of a signal peptide of 24 amino acids, a catalytic region of 634 amino acids exhibiting 50.1% identity with the endo-β-N-acetylglucosaminidase from Arthrobacter protophormiae (Endo-A), and a C-terminal tail of 220 amino acids. Transformed Escherichia coli cells carrying the Endo-BH gene exhibited endo-β-N-acetylglucosaminidase activity. Recombinant Endo-BH hydrolyzed high-mannose type oligosaccharides and hybrid type oligosaccharides, and showed transglycosylation activity. On deletion of 219 C-terminal amino acid residues of Endo-BH, the wild type level of activity was retained, whereas with deletions of the Endo-A homolog domain, the proteins were expressed as inclusion bodies and these activities were reduced. These results suggest that the enzymatic properties of Endo-BH are similar to those of Endo-A, and that the C-terminal tail does not affect the enzyme activity. Although the C-terminal tail region is not essential for enzyme activity, the sequence is also conserved among endo-β-N-acetylglucosaminidases of various origins.
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Dietary Branched-chain Amino Acids Suppress the Expression of Pancreatic Amylase
mRNA in Rats
Naoto Hashimoto and Hiroshi Hara†
Division of Applied Bioscience, Graduate School of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kita-ku, Sapporo 060-8589, Japan
Regulators for pancreatic amylase were examined. Rats were fed ad libitum a 20% amino acid (AA) mixture diet (Con), a 60% AA diet (HA), a branched-chain amino acid (BCAA)-rich diet (BC), or a diet supplemented with AA other than BCAA (OA) for 7 d, or fed the Con, HA, BC diets or diets supplemented with individual BCAA. Activity and mRNA levels of pancreatic amylase in the BC and HA groups were lower than those in the Con and OA groups. Leucine and isoleucine contributed to these effects of the BC diet. The mRNA levels correlated with individual pancreatic BCAA concentrations but not with plasma insulin level. In conclusion, dietary BCAA, especially leucine and isoleucine, may reduce amylase mRNA and activity in rats.
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Enzymatic Properties and Nucleotide and Amino Acid Sequences of a Thermostable
β-Agarase from the Novel Marine Isolate, JAMB-A94
Yukari Ohta, Yuichi Nogi, Masayuki Miyazaki, Zhijun Li, Yuji Hatada, Susumu Ito,† and Koki Horikoshi
Japan Agency for Marine-Earth Science and Technology (JAMSTEC), 2-15 Natsushima, Yokosuka 237-0061, Japan
A gene, agaA, for a novel β-agarase from the marine bacterium JAMB-A94 was cloned and sequenced. The 16S rDNA of the isolate had the closest match, of only 94.8% homology, with that from Microbulbifer salipaludis JCM11542T. The agaA gene encoded a protein with a calculated molecular mass of 48,203 Da. The deduced amino acid sequence showed 37–66% identity to those of known agarases in glycoside hydrolase family 16. A carbohydrate-binding module-like amino acid sequence was found in the C-terminal region. The recombinant enzyme was hyper-produced extracellularly when Bacillus subtilis was used as a host. The purified enzyme was an endo-type β-agarase, yielding neoagarotetraose as the main final product. It was very thermostable up to 60 °C. The optimal pH and temperature for activity were around 7.0 and 55 °C respectively. The activity was not inhibited by EDTA (up to 100 mM) and sodium dodecyl sulfate (up to 30 mM).
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Purification, Cloning, and Sequence Analysis of β-N-Acetylglucosaminidase
from the Chitinolytic Bacterium Aeromonas hydrophila Strain SUWA-9
Xiqian Lan, Naomi Ozawa, Naohide Nishiwaki, Ritsuko Kodaira, Mitsuo Okazaki, and Makoto Shimosaka†
Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, 3-15-1 Tokida, Ueda, Nagano 386-8567, Japan
A chitinolytic bacterium was isolated from Lake Suwa and identified as Aeromonas hydrophila strain SUWA-9. The strain grew well on a synthetic medium containing colloidal chitin as sole carbon source. Chitin-degrading activity was induced by colloidal chitin or N-acetylglucosamine (GlcNAc). Most of the activity, however, was not detected in culture fluid but was associated with cells. A β-N-acetylglucosaminidase was purified after it was solubilized from cells by sonication. The purified enzyme hydrolyzed N-acetylchitooligomers from dimer to pentamer and produced GlcNAc as a final product. The enzyme also hydrolyzed synthetic substrates such as p-nitrophenyl (pNP)-N-acetyl-β-D-glucosaminide and pNP-N-acetyl-β-D-galactosaminide. A gene coding for the purified β-N-acetylglucosaminidase was isolated. The ORF identified is 2,661 nucleotides long and encodes a precursor protein of 887 amino acids including a signal peptide of 22 amino acid residues. The amino acid sequence deduced showed a high similarity to those of bacterial β-N-acetylhexosaminidases classified in family 20 of glycosyl hydrolases.
-14-
Gelling Properties of Soybean β-Conglycinin Having Different Subunit Compositions
Mohamad ramlan Bin Mohamed Salleh,1,2 Nobuyuki Maruyama,1 Koji Takahashi,3 Kazuhiro Yagasaki,4 Takahiko Higasa,1 Yasuki Matsumura,1 and Shigeru Utsumi1,†
1Laboratory of Food Quality Design and Development, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan
2Department of Biotechnology Engineering, Kulliyyah of Engineering, International Islamic University Malaysia, Jalan Gombak 53100, Kuala Lumpur, Malaysia
3National Institute of Crop Science, Kannondai 2-1-18, Tsukuba, Ibaraki 305-8518, Japan
4Nagano Chushin Agricultural Experiment Station, Shiojiri, Nagano 399-6461, Japan
The effects of protein concentration, and heating temperature and time on the gelling properties of soybean β-conglycinin (7S globulins) lacking the α or α’ subunit were compared with those of 7S containing all three subunits (α, α’, and β) to determine whether each subunit contributes equally. In most of the conditions, the relative order of gel hardness was α’-lacking > 7S > α-lacking. From Fourier transform infrared studies, the secondary structure change after heating was very similar among the three samples; thus, the secondary structural change is not the reason for the differences in gel hardness. By using scanning electron microscopy, we observed differences in strand thickness and the density of the gel network among the three samples. These differences correlated well with the differences in gel hardness.
-15-
Synthesis of Brassinosteroids of Varying Acyl Side Chains and Evaluation of
Their Brassinolide-like Activity
Shinya Uesusuki, Bunta Watanabe, Shuji Yamamoto, Junko Otsuki, Yoshiaki Nakagawa,† and Hisashi Miyagawa
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan
Brassinosteroids containing various side chain moieties were synthesized and their activity was determined as the reciprocal logarithm of the ED50 (50% effective dose per plant in moles) in the rice lamina inclination assay using synergist indole-3-acetic acid (IAA). The introduction of a hydroxyl group in the α-position to the carbonyl group of the ester structure significantly enhanced the activity. 2α,3α-Dihydroxy-17β-[(2R,3S)-2-hydroxy-3-methylpentanoyl]oxy-B-homo-7-oxa-5α-androstan-6-one showed the highest activity, for which the pED50 was determined to be 10.5 under synergistic conditions with IAA. Under identical conditions, the pED50 values of brassinolide and castasterone were determined to be 13.6 and 12.3 respectively. With respect to the α-carbon of the acyl moiety, the R-form was 10 times more potent than the corresponding S-form. Substituting the terminal structure (Et) of the side chain to that of the most potent compound, brassinolide (i-Pr), did not increase the activity.
-16-
Isolation of Flavohemoglobin from the Actinomycete Streptomyces antibioticus
Grown without External Nitric Oxide Stress
Yasuyuki Sasaki,1 Naoki Takaya,1 Akira Nakamura,1 and Hirofumi Shoun2,†
1Institute of Applied Biochemistry, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan
2Department of Biotechnology, Graduate School of Agricultural and Life Sciences, University of Tokyo, Bunkyo-Ku, Tokyo 113-8657, Japan
A flavocytochrome protein was isolated from the actinomycete Streptomyces antibioticus. The purified protein contained protoheme and FAD, and its Mr was estimated to be 52,000. The absorption spectra in its resting oxidized, dithionite-reduced, carbon monoxide-bound, and oxygenated (O2-bound) forms were characteristic of those of flavohemoglobin (Fhb). The N-terminal amino acid sequence showed high identities to those of other Fhb’s. Furthermore, the actinomycete flavocytochrome scavenged nitric oxide in the presence of NADH. These results demonstrated that the flavocytochrome is the first Fhb purified from actinomycetes. The actinomycete Fhb was produced in S. antibioticus cells in large amounts without any external nitric oxide (NO) stress, which is indicative of a physiological function of Fhb other than detoxification of NO.
-17-
Effects of Truncation at the Non-homologous Region of a Family 3 β-Glucosidase
from Agrobacterium tumefaciens
Li Ying,1,2 Motomitsu Kitaoka,1,† and Kiyoshi Hayashi1
1Enzyme Laboratory, National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan
2College of Biological Sciences, China Agricultural University, Beijing, 100094, China
The function of the non-homologous region of a family 3 β-glucosidase from Agrobacterium tumefaciens (Cbg1) was studied by analyzing the properties of mutant enzymes that have internal truncated amino acid sequences in the region. Five truncated mutants named Cbg1-d4, Cbg1-d31, Cbg1-d62, Cbg1-d89, and Cbg1-d119 having deletions of 4, 31, 62, 89, and 119 amino acid residues starting from Phe417, respectively, were expressed in Escherichia coli and purified. All the mutants exhibited β-glucosidase activity, indicating that the non-homologous region was not essential for the activity. The truncation caused thermal instability, decrease in pKa of the proton donor residue (Glu616), and deficient transglycosylation activity. The thermal stability and the pKa of Glu616 were partially recovered with longer truncation, suggesting that the truncation perturbed the structure and that their presence in the region was not essential. The main role of the non-homologous region could be formation of a hydrophobic atmosphere at the acceptor site to make the enzyme suitable for hydrolyzing hydrophobic glucosides.
-18-
Identification of High Molecular Weight Proteins in Squid Muscle by Western
Blotting Analysis and Postmortem Rheological Changes
Chinatsu Kasamatsu,1,2,† Sumiko Kimura,3 Mieko Kagawa,4 and Keiko Hatae1
1School of Human Life and Environmental Science, Ochanomizu University, Otsuka 2-1-1, Bunkyo-ku, Tokyo 112-8610, Japan
2Department of Nutrition and Dietetics, Kamakura Women’s University, Ofuna 6-1-3, Kamakura-shi, Kanagawa 247-8512, Japan
3Faculty of Science, Chiba University, Yayoi-cho 1-33, Inage-ku, Chiba 263-8522, Japan
4Komatsu High School, Komatsu-cho Shinyashiki 42-1, Shuso-gun, Ehime 799-1101, Japan
The high molecular weight protein connectin (also called titin) in Japanese common squid (Todarodes pacificus) mantle muscle was identified by western blotting analysis with 3B9, the mouse anti-chicken skeletal muscle connectin monoclonal antibody. Similarly to vertebrate samples, there exists connectin in invertebrate squid mantle muscle, and the amino acid sequences are assumed to resemble those present in the A band of vertebrate connectin, judging by the specificity of 3B9. Moreover, the connectin in squid muscle migrated in this study as a closely spaced doublet of α and β (titins 1 and 2). Between 5 and 7 h post-mortem, the SDS PAGE patterns of the squid sample indicated a change of the doublet bands into a single β-connectin band. Simultaneously, the rheological properties of the squid muscle changed substantially. This degradation of α-connectin into β-connectin in the muscle can explain the critical change that occurs during the post-mortem tenderization of squid muscle.
-19-
Novel Fusicoccins R and S, and the Fusicoccin S Aglycon (Phomopsiol) from Phomopsis
amygdali Niigata 2-A, and Their Seed Germination-stimulating Activity in the
Presence of Abscisic Acid
Naoto Tajima,1 Manabu Nukina,1,2 Nobuo Kato,3 and Takeshi Sassa1,2,†
1Course of the Science of Bioresources, The United Graduate School of Agricultural Sciences, Iwate University (Yamagata University), Ueda-cho, Morioka 020-8550, Japan
2Department of Bioresource Engineering, Faculty of Agriculture, Yamagata University, Wakaba-cho, Tsuruoka 997-8555, Japan
3Institute of Scientific and Industrial Research, Osaka University, Mihogaoka-cho, Ibaraki 567-0047, Japan
Our search for new 3-hydroxyfusicoccins structurally related to cotylenin A from a culture of Phomopsis amygdali Niigata 2-A resulted in the isolation of novel 3-hydroxy fusicoccins, called fusicoccins R and S, and the fusicoccin S aglycon, called phomopsiol, together with known 3α-hydroxyfusicoccin J. The structure of phomopsiol was identified as that of O-demethyl-3-epicotylenol based on spectroscopic evidence. The structures of fusicoccins R and S were also determined to be those of 3’-deacetyl-3α-hydroxyfusicoccin A and 3β-hydroxy-3-epifusicoccin H. The lettuce seed germination-stimulating activity of fusicoccins R and S, phomopsiol and 3α-hydroxyfusicoccin J was examined in the presence of ABA; fusicoccin R and 3α-hydroxyfusicoccin J were highly active, while fusicoccin S and phomopsiol were inactive. The possible biosynthetic relationships among these novel fusicoccins having a 3α- or 3β-hydroxy group in their diterpene moiety are briefly discussed.
-20-
Note
New Monoterpene Glucoside from the Aerial Parts of Thyme (Thymus vulgaris L.)
Hiroyuki Takeuchi,1,† Zhan-Guo Lu,1 and Tomoyuki Fujita2
1Research and Development Laboratories, Nippon Terpene Chemicals, Inc., 4-10 Wakinohama-cho 1-chome, Chuo-ku, Kobe 651-0072, Japan
2Division of Applied Biological Chemistry, Graduate School of Agriculture and Biological Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Sakai, Osaka 599-8531, Japan
A new monoterpene glucoside (1) was isolated from a methanol extract of the dried aerial parts of thyme (Thymus vulgaris L.), together with known 2- and 5-β-D-glucopyranosylthymoquinols (2 and 3, respectively), and (-)-angelicoidenol-β-D-glucopyranoside (4). The structure of 1 was elucidated to be (R)-p-cymen-9-yl β-D-glucopyranoside by spectral evidence and enantioselective synthesis from (R)- and (S)-p-cymen-9-ol derived from p-cymen-8-ol.
-21-
Note
Suppressive Effects of Dietary Fiber in Yogurt on the Postprandial Serum Lipid
Levels in Healthy Adult Male Volunteers
Shizuki Kondo, Jin-zhong Xiao,† Noritoshi Takahashi, Kazuhiro Miyaji, Keiji Iwatsuki, and Sadayuki Kokubo
Food Research and Development Laboratory, Morinaga Milk Industry Co., Ltd., Zama 228-8583, Japan
This study assessed the effect of partially hydrolyzed guar gum (PHGG) in yogurt on the elevation of postprandial serum lipid levels. Eleven healthy adult male subjects were given yogurt with or without 6 g of PHGG in a fat tolerance test as a crossover study. Supplementation with 6 g of PHGG significantly suppressed the incremental peaks and areas under the incremental curve (AUIC) of postprandial serum remnant-like lipoprotein particle cholesterol (RLP-C) and triglyceride (TG). The results suggest the potential of PHGG to reduce the risk of hyperlipemia.
-22-
Note
Design of Soymetide-4 Derivatives to Potentiate the Anti-alopecia Effect
Takahiro Tsuruki and Masaaki Yoshikawa†
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011, Japan
Previously, we found that soymetide-4 (MITL), an N-formyl-methionyl-leucyl-phenylalanine (fMLP) agonist peptide derived from soybean β -conglycinin α ’ subunit, stimulated phagocytosis of human polymorphonuclear leukocytes, and inhibited alopecia induced by etoposide, an anticancer drug, in neonatal rats after oral administration. We found that the fMLP receptor affinity and phagocytosis-stimulating activity of soymetide-4 was potentiated by replacement of Thr3 with hydrophobic residues. Among the derivatives synthesized, [Trp]3-soymetide-4 (MIWL) was the most potent, stronger by 180 and 130 times than soymetide-4 in receptor affinity and phagocytosis-stimulating activity, respectively. The anti-alopecia effect of [Trp]3-soymetide-4 was about 3 times larger than that of soymetide-4 after oral administration.
-23-
Note
Enzymatic Reactions by Five Chalcone Synthase Homologs from Hop (Humulus lupulus
L.)
Yukio Okada,1,† Yukie Sano,2 Takafumi Kaneko,1 Ikuro Abe,2 Hiroshi Noguchi,2 and Kazutoshi Ito1
1Bioresources Research and Development Laboratories, Sapporo Breweries, Ltd., 37-1 Kizaki, Nitta, Gunma 370-0393, Japan
2School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Shizuoka 422-8526, Japan
The enzyme activities encoded in five cDNAs for chalcone synthase (CHS) homologs from hop were investigated. Only valerophenone synthase (VPS) and CHS_H1 showed both naringenin-chalcone and phlorisovalerophenone forming activity. Narigenin-chalcone production by VPS was much lower than by CHS_H1. Therefore, it is highly possible that flavonoid depends mainly on CHS_H1, while bitter acid biosynthesis depends mainly on VPS and CHS_H1.
-24-
Note
Isolation of an Exopolysaccharide-producing Bacterium, Sphingomonas sp. CS101,
Which Forms an Unusual Type of Sphingan
Eun-Jung Seo,1,* Sang-Ho Yoo,2,* Ko-Woon Oh,1 Jaeho Cha,3 Hyeon Gyu Lee,4 and Cheon-Seok Park1,†
1Department of Food Science and Technology, Kyunghee University, Yongin 449-701, Korea
2Department of Food Science and Technology, Sejong University, Seoul 143-747, Korea
3Department of Microbiology, Pusan National University, Busan 609-735, Korea
4Department of Food Science and Nutrition, Hanyang University, Seoul 133-791, Korea
An exopolysaccharide-producing Gram negative bacterium was isolated and determined to be a Sphingomonas sp. (CS101). A sugar composition analysis of an exopolysaccharide indicated that the Sphingomonas sp. CS101 secreted an exopolysaccharide composed of glucose, mannose, fucose, and rhamnose in the ratio of 2.1:1.1:1.0:0.1, suggesting that this exoplysaccharide is an unusual type of sphingan family. The mean molecular weight of the exopolysaccharide was determined to be 4.2× 105 Da by size exclusion chromatography coupled with multi-angle laser-light scattering (SEC/MALLS) analysis. An exopolysaccharide was produced up to 17 g/l (pH 7; 30 °C) with the optimal medium condition over 4 days of cultivation.
-25-
Note
Catalytic Activity of Tripeptidase from Lactococcus lactis to Which Amino Acid
Substitution Was Introduced According to Natural Mutation
Sumiko Mori, Satoshi Kaneko, and Takafumi Kasumi†
National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki 305-8642, Japan
Four mutations observed between tripeptidases from Lactococcus lactis subsp. lactis and subsp. cremoris were introduced one by one to the corresponding points in wild-type tripeptidase from L. lactis subsp. lactis. The kcat values of four resultant mutants were analyzed and discussed in stereographical terms. Change in catalytic activity appeared to be related to the sequential and steric location of mutation point within the enzyme protein, even though no drastic change was observed with one point mutation.
-26-
Note
Effects of Soybean β -Conglycinin on Hepatic Lipid Metabolism and Fecal
Lipid Excretion in Normal Adult Rats
Kensuke Fukui,1,† Makiko Kojima,1 Nobuhiko Tachibana,1 Mitsutaka Kohno,1 Kiyoharu Takamatsu,1 Motohiko Hirotsuka,1 and Makoto Kito2
1Food Science Research Institute, Fuji Oil Co., Ltd., 1 Sumiyoshi-cho, Izumisano, Osaka 598-8540, Japan
2Emeritus Professor of Kyoto University
β -Conglycinin decreased blood triacylglycerol (TAG) levels in male Wistar adult rats. Liver mitochondrial carnitine palmitoyltransferase activity in the β -conglycinin-fed group significantly increased as against the casein-fed group. Hepatic fatty acid synthase activity in the β -conglycinin group significantly decreased as against that of the casein-fed group. Fecal fatty acid excretion in the β -conglycinin group was significantly higher than in the casein group.
-27-
Note
Involvement of a Glu71–Arg64 Couple in the Access Channel for NADH in
Cytochrome P450nor
Fei Su,1 Shinya Fushinobu,2 Naoki Takaya,1 and Hirofumi Shoun2,†
1Institute of Applied Biochemistry, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan
2Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan
Putative access channel for NADH in the heme-distal pocket of cytochrome P450nor (P450nor) comprises many charged amino acid residues. Characterization of the E71A mutant protein of P450nor highlights the existence of a unique mechanism for binding NADH that depends on the salt bridge network between Glu71, Arg64 and Asp88.
-28-
Note
Resistance Imparted by Traditional Chinese Medicines to the Acute Change of
Glutamic Pyruvic Transaminase, Alkaline Phosphatase and Creatine Kinase Activities
in Rat Blood Caused by Noise
Bei-Wei Zhu,1 Yu-Mei Sun,1 Xia Yun,1 Song Han,2 Mei-Lan Piao,2 Yoshiyuki Murata,3 and Mikiro Tada2,†
1College of Bio & Food Technology, Dalian Institute of Light Industry, Dalian 116034, China
2Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan
3Department of Agriculture, Okayama University, Okayama 700-8530, Japan
The activities of serum glutamic pyruvic transaminase (GPT), alkaline phosphatase (ALP) and creatine kinase (CK) in rats injected or not with the Chinese medicines, Astragali, Rhodiolae and Ligusticum, were determined after noise exposure. Noise at 95 and 105 dB significantly increased the activities of GPT, ALP and CK, and showed a dependence on the exposure time. The injection of each medicine significantly suppressed the increased enzyme activities by 95 and 105 dB noise.
-29-
Note
Biotransformation of α-Isomethylionone to 1-(2,6,6-Trimethyl-2-cyclohexen-1-yl)propan-2-one
Susumu Ishizaki,1 Masamichi Itoh,1,† Tsuyoshi Komai,1 Tsutomu Honda,1 and Takeshi Kitahara2
1Technical Research Center, T. Hasegawa Co., Ltd., 335 Kariyado, Nakahara-ku, Kawasaki 211-0022, Japan
2Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan
The main biodegradation product of (±)-α-isomethylionone (2) with standard activated sludge was characterized as (±)-1-(2,6,6-trimethyl-2-cyclohexen-1-yl)propan-2-one (1) by its analysis and synthesis. Both enantiomers (1a and 1b) of 1 were synthesized by starting from (R)- and (S)-2,4,4-trimethyl-2-cyclohexen-1-ol (3a and 3b), respectively.
-30-
Note
Microbial Hydroxylation of Indole to 7-Hydroxyindole by Acinetobacter calcoaceticus
Strain 4-1-5
Daisuke Sugimori,1,† Takanori Sekiguchi,1 Fumihiko Hasumi,2 Motoki Kubo,3 Naoki Shirasaka,4 and Masaya Ikunaka4
1Department of Chemistry and Biology Engineering, Fukui National College of Technology, Geshi, Sabae, Fukui 916-8507, Japan
2Department of Chemistry and Biochemistry, Numazu College of Technology, 3600 Ooka, Numazu, Shizuoka 410-8501, Japan
3Department of Bioscience and Biotechnology, Faculty of Science and Engineering, Ritsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga 525-8577, Japan
4Research & Development Center, Nagase & Co., Ltd., 2-2-3 Murotani, Nishi-ku, Kobe, Hyogo 651-2241, Japan
A screening study yielded Acinetobacter calcoaceticus strain 4-1-5, which is capable of hydroxylating indole to 7-hydroxyindole. Strain 4-1-5 grew on terephthalate as the sole source of carbon and energy and hydroxylated indole to 7-hydroxyindole by cometabolism of indole using terephthalate as cosubstrate. Strain 4-1-5 produced 0.574 mM of 7-hydroxyindole at 2.38 mM indole in 24 h with the cell growth.
-31-
Communication
Characterization of Protein Phosphatase 2A Acting on Phosphorylated Plasma Membrane
Aquaporin of Tulip Petals
Abul Kalam Azad,1,2 Yoshihiro Sawa,1 Takahiro Ishikawa,1 and Hitoshi Shibata1,†
1Faculty of Life and Environmental Sciences, Shimane University, Matsue, Shimane 690-8504, Japan
2The United Graduate School of Agricultural Sciences, Tottori University, Koyama-Minami, Tottori 680-0945, Japan
A protein phosphatase holo-type enzyme (38, 65, and 75 kDa) preparation and a free catalytic subunit (38 kDa) purified from tulip petals were characterized as protein phosphatase 2A (PP2A) by immunological and biochemical approaches. The plasma membrane containing the putative plasma membrane aquaporin (PM-AQP) was prepared from tulip petals, phosphorylated in vitro, and used as the substrate for both of the purified PP2A preparations. Although both preparations dephosphorylated the phosphorylated PM-AQP at 20 °C, only the holo-type enzyme preparation acted at 5 °C on the phosphorylated PM-AQP with higher substrate specificity, suggesting that regulatory subunits are required for low temperature-dependent dephosphorylation of PM-AQP in tulip petals.