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Volume 68 (1970)
- Issue 6 Pages 763-
- Issue 5 Pages 603-
- Issue 4 Pages 427-
- Issue 3 Pages 255-
- Issue 2 Pages 137-
- Issue 1 Pages 1-
Volume 68, Issue 4
Displaying 1-21 of 21 articles from this issue
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Michihiko OGUCHI1970 Volume 68 Issue 4 Pages 427-439
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe physical and catalytic properties of crystalline glyceraldehyde-3-phosphate dehy-drogenase [EC 1. 2. 1. 12] from human erythrocytes were compared with those of the enzyme from rabbit muscle. The molecular weight of the erythrocyte enzyme was found to be 137, 000 which was close to that of the rabbit muscle enzyme. The two enzymes behaved differently on electrophoresis. The amino acid compositions of the two enzymes were very similar though there were eight half cystines per mole of the erythrocyte enzyme: half as many as in the rabbit enzyme. Four of the sulf-hydryl groups in the enzyme were apparently free and were essential for enzyme activity. The other four SH-groups were hidden but reacted with p-chloromercuri-benzoate when the enzyme was denatured with urea. The erythrocyte enzyme con-tained less than one mole of NAD, which was bound firmly to the crystallized prepa-ration. However, it was found that the enzyme could bind approximately 3. 5 moles of the coenzyme. The enzyme from erythrocytes had essentially the same molec-ular activity, pH-activity curve and substrate specificity as the rabbit enzyme. Kinetic studies indicated that the reaction of the erythrocyte enzyme was an ordered mechanism of substrate addition. Various kinetic parameters of the reaction are reported.View full abstractDownload PDF (2087K) -
Tomoko YAMAGUCHI1970 Volume 68 Issue 4 Pages 441-447
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSBilirubin inhibition of crystalline beef liver glutamate dehydrogenase [EC 1. 4. 1.3] was demonstrated. Bilirubin glucuronide was a more potent inhibitor than free bulirubin, and albumin bound bilirubin had the least inhibitory effect on the enzyme. Bilirubin glucuronide was possibly contaminated by bile salts, which may have ac-celerated the inhibitory effect. Bilirubin inhibition was reversible in the presence of zinc ion at 2×10-2M under the present experimental conditions. Three modes of inhibition by bilirubin were suggested; 1) zinc chelation, 2) denaturation by deter-gent action, and 3) competition with the pyridinium ring of NAD at the binding site.View full abstractDownload PDF (544K) -
Masaaki HIROSE, Etsuro SUGIMOTO, Ryuzo SASAKI, Hideo CHIBA1970 Volume 68 Issue 4 Pages 449-457
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA procedure for crystallization of phosphoglucomutase [EC 2. 7. 5. 1] from baker's yeast is presented. The purified enzyme preparation was homogeneous as judged from ultracentrifugation and electrophoresis.
Yeast phosphoglucomutase was not influenced by the addition of Mg2+ and was strongly inhibited by chelating-agents. However, the enzyme was stimulated about 1.7-fold by simultaneous addition of Mg2+ and chelating-agents without preincubation.
The reaction mechanism of the yeast enzyme was studied. It was concluded by kinetic experiments that the enzyme-substrate-coenzyme ternary complex is the active intermediate in yeast phosphoglucomutase reaction. It was suggested in the pattern of Lineweaver-Burk plots that the reaction mechanism in yeast phosphoglu-comutase fitted Cleland's “random sequential” pathway and substrate binding to the enzyme decreased the affinity of coenzyme to the enzyme, and vice versa. This mechanism was able to interpret also the phenomena in the rabbit muscle enzyme supporting “ping-pong” pathway.View full abstractDownload PDF (2696K) -
Keizo WAKU, Yasuo NAKAZAWA1970 Volume 68 Issue 4 Pages 459-466
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. The activity acylating 1-O-alkyl-glycero-3-phosphorylcholine was found for the first time in rabbit sarcoplasmic reticulum, which also exhibits acyl transfer activities to other phosphoglycerides. The product of the enzymatic reaction was proved to be 1-O-alkyl-2-acyl-glycero-3-phosphorylcholine.
2. The acyl transfer activity to 1-O-alkyl-glycero-3-phosphorylcholine was estimated to be significantly lower than that to 1-acyl-and 1-O-alkenyl analogues in the same tissue.
3. The acyl transfer rates with different CoA thiol esters were highly specific to poly-unsaturated fatty acyl-CoA in contrast to acyl-CoA: 1-acyl-analogue acyltrans-ferase having activity to both saturated and unsaturated derivatives.View full abstractDownload PDF (639K) -
I. Immunological Memory Cells Separated from Spleen and Lymph Nodes of the Immunized RabbitsMunehiko YUKIOKA, Yoshihiro KINOSHITA, Tadashi TAKESHITA, Eiichi KIMUR ...1970 Volume 68 Issue 4 Pages 467-473
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe lymphoid cells prepared from spleen or lymph nodes of immunized rabbits were fractionated into three groups by multi-layer centrifugation. The most dense cellular population which contained the highest percentage of small lymphocyte showed the specific enhancement of DNA synthesis by the anamnestic antigenic stimulation. During secondary immune response, the immunological memory cells seemed to be converted to a less dense type, and antibody was produced mainly in these less dense cells.
Various kinds of antimetabolite of nucleic acid and protein synthesis have shown the remarkable inhibitory effects on antibody formation in vitro system.View full abstractDownload PDF (1168K) -
Tamotsu TAKETOMI, Nariko KAWAMURA1970 Volume 68 Issue 4 Pages 475-485
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDeacylated hematoside prepared from hernatoside of equine erythrocyte by a partial alkaline hydrolysis was purified by Silica Gel H-Hyfio Super Cel column chromatog-raphy and its homogeneity was detected by a thin layer chromatogram showing one single spot. The chemical structure of the deacylated hematoside was found to be neuraminyl (2→3) galactosyl (1→4) glucosylsphingosine by the following procedures: chemical analysis of each component of the sample, gas-liquid chromatography of methyl glycosides obtained after methanolysis of the permethylated one and forma-tion of lactosyl sphingosine from the sample by mild acid hydrolysis releasing a neuraminic acid.
Hemolytic activity of the deacylated hematoside was about twice stronger than that of psychosine, sphingosylphosphorylcholine, lysolecithin et al. as a calculated L50 value per red cell. Thus the deacylated hematoside may be reasonably called “lysohematoside” belonging to the so called “lysosphingolipids. ” It was found that the lysosphingolipids including the lysohematoside did not produce any O-methyl-sphingosine as artifact by anhydrous methanolysis contrary to sphingolipids. A hy-pothetical mechanism of the formation of O-methyl-sphingosine from sphingolipids by the anhydrous methanolysis was suggested.View full abstractDownload PDF (2218K) -
Akio YOSHIMOTO, Hiroyuki ITO, Kenkichi TOMITA1970 Volume 68 Issue 4 Pages 487-499
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effects of various cofactors on prostaglandin formation from arachidonic acid by the microsomal enzyme of bovine seminal vesicles were studied. In addition to the known activators, reduced glutathione and hydroquinone, the enzyme system was found to be markedly stimulated by hemoglobin, myoglobin or hemin.
From the specific effects of these cofactors on prostaglandin formation or oxygen uptake, or both, it seemed that heme compounds and hydroquinone were involved in the step in which molecular oxygen became attached to the unsaturated fatty acids, while reduced glutathione was involved in the subsequent step of reduction of the peroxide-type intermediate.
The facts that hydroquinone could be replaced by ascorbate, and that the order of addition of reactants to the system markedly affected both the yield of prostaglandin and the rate of oxygen consumption suggested that a free radical was formed during prostaglandin biosynthesis.View full abstractDownload PDF (1034K) -
Kazuko ANDO1970 Volume 68 Issue 4 Pages 501-508
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Conditions for preparation of apo-laccase [EC 1. 10. 3. 2] and reconstruction of the holo-enzyme by treatment with copper ion were examined. The apo-enzyme was prepared by dialysis against 10-20mM cyanide solution containing 20-50mM ascorbate at pH 7.0-7.4. The properties of native laccase, such as its activity, copper content, absorption spectrum and electron spin resonance spectrum, were almost completely regained by treatment of this apo-protein with cuprous ion and the percentage re-construction was 70 to 90%. Some, though much less reconstruction was achieved by treatment of apo-enzyme with cupric ion.
2. The relationships between the copper content, the absorbancy at 615mμ and the activity were studied using “partial apo-enzymes, ” derivatives of native enzyme containing various amounts of copper obtained during cyanide-treatment. The de-crease in activity of the “partial apo-enzymes” was much larger than the decrease in their copper content. Thus, removal of half the initial amount of copper resulted in loss of more than 90% of the original activity. The cooperative action of copper atoms in the laccase reaction is suggested.View full abstractDownload PDF (587K) -
Akira KOTAKI, Kunio YAGI1970 Volume 68 Issue 4 Pages 509-516
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe fluorescence properties of riboflavin derivatives in various media have been studied by use of a sensitive apparatus that renders the direct measurement of cor-rected emission spectra in the wavelength range of 340-660mμ. The corrected emis-sion (quantum) and excitation spectra of riboflavin tetrabutyrate in n-heptane have been recorded since it is slightly soluble in this hydrocarbon. The quantum spectrum shows clear peaks at 493 and 522mμ and a shoulder at 560mμ, thus giving apparent mirror image to the three-banded longest wavelength band of the corrected excita-tion spectrum which is expected to coincide with the absorption spectrum of this compound in n-heptane. General features of flavin fluorescence observed by chang-ing the solvent from apolar to polar type are the red-shift of the emission maximum, the broadening of the peak, the disappearance of the three-banded structure, and the reduction of radiation probability. In particular, in the aqueous binary mixtures, the emission maximum and quantum yield of flavin fluorescence vary as a function of dielectric constant or of empirical Z-value that reflects the polarity of solvents. However, such linear correlation has not always been found when the effects of pure solvents of wide variety are tested. This fact suggests that the fluorescence proper-ties of flavin are also considerably influenced by the solvent characteristics other than dielectric constants.View full abstractDownload PDF (609K) -
X. Reactions of Iodoacetate, Iodoacetamide and Related Alkylating Reagents with Ribonuclease T1Kenji TAKAHASHI1970 Volume 68 Issue 4 Pages 517-527
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. The alkylation of the γ-carboxyl group of glutamic acid-58 was shown to be the predominant reaction and the major cause of inactivation in the reaction of iodoacetate with ribonuclease T1 [EC 2. 7. 7. 26] at pH 7.5 and 8.5 as well as at pH 5.5. At pH 7.5 and 8.5, however, histidine residues were also alkylated, though the rate was much slower. The pH dependency of the rate of inactivation with iodoacetate in-dicated implication of two groups in the reaction with a pK value of about 4 (pos-sibly glutamic acid-58) and about 7 (histidine residue(s)).
2. Glutamic acid-58 was alkylated at both pH 5.5 and pH 8.0 by a number of mono-and dibromo acids with concomitant inactivation of the enzyme. Among the reagents studied, β-bromopyruvate inactivated the enzyme nearly as rapidly as iodo (or bromo) acetate, but other reagents were much less reactive. The reactivity of bromo acids decreased greatly with increasing the size of the substituent on the carbon atom to which the bromine atom is attached.
3. The reaction of iodoacetamide and the related reagents having a bromo (or iodo) acetamido group was slower and less specific than that of iodoacetate. Unlike iodo-acetate, these reagents inactivated the enzyme more rapidly at pH 8.0 than at pH 5.5. The reaction seemed to involve some histidine and the amino-terminal alanine res-idues. Alkylation reaction may occur also with glutamic acid-58 depending on the reaction condition employed. The reactions of these reagents also suggested that two of the three histidine residues in the enzyme molecule are preferentially alkylated by these reagents and that one or both of these two are essential for the enzymatic activity. The third histidine residue appears to be buried in the interior of the protein molecule or bound to some other residues, thus becoming much less reactive. Otherwise, there may be two histidine residues which are alkylated only in a mu-tually exclusive way.View full abstractDownload PDF (877K) -
VII. Reconstitution of a Steroid 11β-Hydroxylation System using a P-450 Preparation Extracted and Partially Purified from Adrenocortical MitochondriaFumiko MITANI, Shigeo HORIE1970 Volume 68 Issue 4 Pages 529-542
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA steroid 11β-hydroxylation system was reconstituted from NADPH-adrenodoxin re-ductase, adrenodoxin, and a P-450 preparation which had been extracted and partially purified from mitochondria of bovine adrenal cortex.
1. Oxidation of NADPH by the reconstituted system was accelerated by addi-tion of deoxycorticosterone, and the formation of corticosterone was detected both chromatographically and fluorometrically. The reductase, adrenodoxin, and the P-450 preparation were all indispensable for the hydroxylation reaction.
2. NADPH oxidation activity in the presence and absence of deoxycorticosterone was measured under various conditions. The results indicated that in the oxidation of N ADPH in the absence of deoxycorticosterone so-called “leakage of electrons” is predominant when the concentration of reduced adrenodoxin was relatively low. Acceleration of the oxidation of NADPH by deoxycorticosterone increases with the concentration of reduced adrenodoxin. “Leakage of electrons” is maximal at pH values around 6.5, whereas acceleration of oxidation by deoxycorticosterone is maxi-mal at about pH 7.5.
3. Of the steroids tested only those inducing a type I difference spectrum of P-450 accelerated the oxidation of NADPH by the reconstituted system. The specificity of the reconstituted system for steroids, as measured by their acceleratory effects, agreed essentially with that of adrenal cortex mitochondria.
4. Pregnenolone, which induced a type lI difference spectrum, inhibited the “leakage of electrons, ” but did not affect the acceleration of NADPH oxidation by deoxycorticosterone.
5. These results are most easily explained by supposing that P-450 existed in two states in the preparation, one of which is responsible for the “leakage” and the other for the hydroxylation reaction.
6. Considerable activity for cleavage of the cholesterol side chain was detected in the reconstituted system using isotopically labelled cholesterol.View full abstractDownload PDF (1139K) -
Shoichi MIURA, Hachiro NAKAGAWA1970 Volume 68 Issue 4 Pages 543-548
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSL-Serine dehydratase [EC 4. 2. 1. 18] activity in the liver of newborn rats developed immediately after birth. Pluse-labeling with 14C-amino acids followed by specific precipitation of the enzyme with antiserum showed that the increase in the enzyme activity was due to net synthesis of enzyme protein.
Serine dehydratase activity in prenatal liver was low and did not change within 8 hr on exposure of the fetus to the external environment by Caesarean operation after 21 days of gestation, while it increased when glucagon was administered in-traperitoneally to the fetus. Induction of serine dehydratase activity by glucagon was due to increase in the rate of synthesis of enzyme molecules.
Insulin repressed prenatal enzyme induction by glucagon and natural postnatal development of liver serine dehydratase. The possibility that glucagon and insulin participate in the enzyme development after birth is discussed.View full abstractDownload PDF (493K) -
Masaki IWABUCHI, Kunihiko ITO, Hiroshi OCHIAI1970 Volume 68 Issue 4 Pages 549-559
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe sedimentation analysis of the ribosomes of Dictyostelium discoideum indicated that the monomeric ribosome had a sedimentation coefficient of about 83 S and could be dissociated into 61-S and 42-S ribosomal subunits. The 83-S ribosome contained 26-S and 17-S rRNA with a mass ratio of approximately 2:1. The 61-S ribosomal subunit contained 26-S rRNA and 4-5-S RNA, whereas the 42-S subunit contained only 17-S rRNA. When these subunits were suspended in 0.1mM Mg2+ for a long period, treated with EDTA, or incubated with 0.5-M KCl, NaCl or LiCl in the pres-ence of 0.1mM Mg2+ for 1 hr, the sedimentation values of both subunits were de-creased to a great extent. The small subunit was more sensitive to RNase [EC 2. 7. 7. 16] digestion than the large subunit. The electrophoretic analysis of basic proteins of ribosomes showed that the 61-S and 42-S subunit contained 21 and 18 components, respectively.View full abstractDownload PDF (1477K) -
Tadayoshi OKUMURA, Ikuo YAMASHINA1970 Volume 68 Issue 4 Pages 561-571
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. α-Mannosidase [EC 3. 2. 1. 24] was purified from hog kidney 1, 200 fold over the crude extract. The purified preparation had a specific activity of 19 units per mg protein when assayed at pH 4.6, the optimum pH, using p-nitrophenyl-α-D-mannoside as substrate, and was free from other glycosidases found in the kidney extract. The enzyme activity of crude preparations was heat stable, resisting a treatment at 60°C for 15min, but the purified enzyme required Zn2+ to retain the activity even at room temperature. The enzyme solution could be stored indefinitely at -20°C without any loss of activity.
2. The purified enzyme was used to study the carbohydrate structures of glyco-peptides from ovalbumin, Taka-amylase [EC 3. 2. 1. 1] and stem bromelain [EC 3. 4. 4. 24]. From determinations of the liberated mannose and analysis of the remaining glycopeptides, aglycon specificity of the enzyme was discussed.View full abstractDownload PDF (804K) -
Kunio YAGI, Nobuhiko SUGIURA, Hirobumi OHAMA, Akira KOTAKI1970 Volume 68 Issue 4 Pages 573-575
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
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Masaru KAWAMURA, Tomoh MASAKI, Yoshiaki NONOMURA, Koscak MARUYAMA1970 Volume 68 Issue 4 Pages 577-580
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
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Shigeru AONUMA, Takao HAMA, Nanaya TAMAKI1970 Volume 68 Issue 4 Pages 581-583
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
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Tadahiko ANDO, Takeo KOSAWA, Yonosuke IKEDA1970 Volume 68 Issue 4 Pages 585-588
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
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Ryuichi KATO, Akira TAKANAKA, Kinichi ONODA, Yoshihito OMORI1970 Volume 68 Issue 4 Pages 589-592
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (241K) -
Tohru KANAZAWA, Shinpei YAMADA, Yuji TONOMURA1970 Volume 68 Issue 4 Pages 593-595
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
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Koichi IWAI, Koei HAMANA, Hiroko YABUKI1970 Volume 68 Issue 4 Pages 597-601
Published: October 25, 1970
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (921K)
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