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Volume 74 (1973)
- Issue 6 Pages 1075-
- Issue 5 Pages 863-
- Issue 4 Pages 639-
- Issue 3 Pages 405-
- Issue 2 Pages 203-
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Volume 74, Issue 6
Displaying 1-26 of 26 articles from this issue
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XIX. Differential Cleavages of the Disulfide Bonds in Ribonuclease T1 with 2-MercaptoethanolSusumu HAYAKAWA, Kenji TAKAHASHI1973 Volume 74 Issue 6 Pages 1075-1081
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. The two disulfide bonds in ribonuclease T1 [EC 2. 7. 7. 26] were reduced by treatment of the enzyme (0.15 or 0.3% solution) with a 1400-fold molar excess of 2-mercaptoethanol at pH 8.8 and 25°C. Under these conditions, about 1.5 disulfide bonds per molecule of protein were reduced in 4hr and the enzymatic activity toward RNA was lost nearly completely. The two disulfide bonds were reduced at different rates; one was reduced much faster than the other, and a considerable extent of inactivation occurred with the reduction of the former.
2. To determine the extent of reduction of each disulfide bond, partially reduced derivatives of ribonuclease T1 were carboxymethylated with iodoacetate, oxidized with performic acid, and hydrolyzed with chymotrypsin. Fractionation by high voltage paper electrophoresis and analyses of the resulting peptides revealed that the disulfide bond between half-cystine residues at positions 2 and 10 was reduced much faster than that between half-cystine residues at positions 6 and 103.View full abstractDownload PDF (518K) -
Kenji TAKAHASHI1973 Volume 74 Issue 6 Pages 1083-1089
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. S-Carboxymethylcysteine was submitted to performic acid oxidation and subsequent acid hydrolysis and the products obtained at each step were analyzed with an amino acid analyzer. Upon oxidation at -10 to -20°C, the major product was S-CM-cysteine sulfoxide (80%), and a small amount of S-CM-cysteine sulfone was also formed. When the oxidation was performed at 37°C, the product was exclusively S-CM-cysteine sulfone.
2. Upon acid hydrolysis, S-CM-cysteine sulfoxide was completely decomposed to give cystine plus cysteine as the major products (47-80%) and S-CM-cysteine as a minor product (maximally 8%). S-CM-cysteine sulfone was also decomposed extensively (maximally 81%) on acid hydrolysis, but gave no ninhydrin-positive amino acid detectable on amino acid analysis.
3. Similar results were obtained with a reduced carboxymethylated derivative of glutathione and of ribonuclease T1 [EC 2. 7. 7. 26] and with reduced carboxamidomethylated ribonuclease A [EC 2. 7. 7. 16]. The yields of cystine plus cysteine from the S-carboxymethyl- or S-carboxamidomethylcysteine residues in these samples upon oxidation and acid hydrolysis were in the range of 40 to 50%.
4. The results obtained with S-CM-cysteine were compared with those obtained with S-methylcysteine and methionine under the same conditions. The sulfoxides of these latter amino acids did not yield cystine and cysteine on acid hydrolysis. The presence of carboxymethyl group on the sulfur atom of S-CM-cysteine sulfoxide appears to be responsible for the preferential cleavage of the C-S bond in the HOOC-CH2-_??_- group of this amino acid over the cleavage of the S-O bond.View full abstractDownload PDF (452K) -
IX. Kinetic Studies on the Conversion of Osmotic Energy to Chemical Energy in the Sarcoplasmic ReticulumShinpei YAMADA, Yuji TONOMURA1973 Volume 74 Issue 6 Pages 1091-1096
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe measured the time-course of P-incorporation from Pi into the SR-ATPase (EP-formation) in medium consisting of 0.1M KCl and 50mM Tris-maleate, pH 7.0 at 5°C, using SR pre-loaded with Ca2+ ions. The amount of EP formed increased linearly with time, after a short lag phase. Therefore, the rate of P-incorporation (EP-formation), vf, was measured from the linear part of the plot of [EP] versus time.
The dependence of the rate of EP-formation, vf, on the concentrations of Mg2+ ions and Pi and the Ca2+-gradient was found to be given by _??_. Under the experimental conditions used, the values of Vf, KP, KMg, and KCa were 1.7moles/106g/sec, 18mM, 9.6mM, and 6.7×108, respectively.View full abstractDownload PDF (357K) -
VII. Interaction of Hydrophobic Probes with D-Amino Acid OxidaseMakoto NAOI, Akira KOTAKI, Kunio YAGI1973 Volume 74 Issue 6 Pages 1097-1105
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. A hydrophobic probe, 4-benzoylamido-4'-aminostilbene-2, 2'-disulfonate (MBAS) was found to fluoresce strongly when bound to the apoenzyrne of D-amino acid oxidase [D-amino acid: O2 oxidoreductase (deaminating), EC 1. 4. 3. 3]. This enhancement of fluorescence was accompanied by a blue shift of the emission maximum of the dye, suggesting that the polarity of the binding site of the apoenzyme is smaller than the polarity of the media. The adduct was found to be an equimolar complex between the apoenzyme and the dye; the dissociation constant for the complex was determined to be 1.47×10-5M. The enhanced fluorescence of MBAS bound to the apoenzyme molecule was completely quenched by the addition of an equimolar amount of FAD, suggesting that the MBAS was expelled from the binding site of the apoenzyme. This was confirmed by an equilibrium dialysis experiment. Kinetic experiments indicated that MBAS binds with the apoenzyme in competition with FAD.
2. Another hydrophobic probe, 1-anilinonaphthalene-8-sulfonate (ANS) also formed an equimolar complex with the apoenzyme of this oxidase, with a considerable enhancement of ANS fluorescence, accompanied by a blue shift in the fluorescence emission maximum. Upon addition of FAD, the enhanced fluorescence was also completely quenched, but liberation of the dye was not observed. The quenching of the fluorescence of the bound ANS caused by the further binding of the apoenzyme with FAD was considered to be due to the interaction between bound ANS and bound FAD, and to the decrease in hydrophobicity of the enzyme locus surrounding the bound ANS molecule. Kinetic experiments indicated that ANS binds with the enzyme in competition with the substrate, D-alanine.View full abstractDownload PDF (622K) -
Michio YAZAWA, Fumi MORITA, Koichi YAGI1973 Volume 74 Issue 6 Pages 1107-1117
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe ultraviolet absorption difference spectrum of heavy meromyosin induced by ATP was followed in the presence of 10-3M MnCl2. The life time of the difference spectrum was almost one second regardless of the initial concentration of ATP. This time was far longer than that of the initial burst of Pi-liberation of ATPase [EC 3. 6. 1. 3]. The difference spectrum after decay was in the shape of an ADP-induced one.
On addition of MgCl2 (final concentration, 10-2M) to heavy meromyosin solution containing 10-4M MnCl2 at a time when an adequate concentration of ATP still remained, the shape of the difference spectrum, which was already in the form of an ADP-induced one, changed immediately to that of an ATP-induced one.
The binding of ADP to heavy meromyosin was measured by the gel-filtration method in the presence of MnCl2. The Scatchard plot was concave upwards and the maximum binding number obtained was 2.1 moles per 3.65×105g of heavy meromyosin. Comparison between the degree of binding and magnitude of the difference spectrum induced by ADP showed that the maximum value of ΔA was attained at one mole of ADP binding.
The maximum value of the initial burst of Pi-liberation was about 1.3 moles per 3.65×105g of heavy meromyosin in the presence of MnCl2. The initial burst decreased through the addition of ADP, without any change in the steady state ATPase rate. Based on these results, the mechanism of ATPase action is discussed.View full abstractDownload PDF (745K) -
Hiroshi HASHIZUME1973 Volume 74 Issue 6 Pages 1119-1130
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIt was found that three of eight tryptophyl residues in α-chymotrypsinogen and π-chymotrypsin [EC 3. 4. 4. 5] were in an exposed state and that one was partially buried at neutral pH. This partially buried tryptophyl residue may play an important role at the substrate-binding site of π-chymotrypsin. Spectrophotometric titration in the acid region, which was essentially parallel to a protonation blue shift, indicated the presence of carboxyl group(s) in contact with this tryptophyl residue. Two-step processes of unfolding around the five buried or partially buried tryptophyl residues were proposed on the basis of heat denaturation studies of the proteins. The partially buried tryptophyl residue was sensitive to heat denaturation, and the four other tryptophyl residues which were completely buried in the proteins, were unfolded at higher temperatures.View full abstractDownload PDF (913K) -
Hideki MIYAJI, Akira MISAKI1973 Volume 74 Issue 6 Pages 1131-1139
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe partially sulfated dextrans (D. S., 0.3-0.77) were prepared from three fractions of acid-degraded dextran of Leuconostoc mesenteroides N-4, by the reaction with either (A) chlorosulfonic acid in formamide or (B) sulfur trioxide-pyridine complex in dimethyl sulfoxide, and the locations of the sulfate groups were investigated by methylation and Smith degradation techniques. Sulfate groups in the sulfated dextrans were distributed rather at random, as glucose mono-, di-, and also trisulfates, although the C-2 position appeared to be most reactive towards sulfation. The results also suggested that the sulfation of dextran by method (B) is more homogeneous than by method (A), so far as intramolecular distribution of the sulfate groups is concerned.View full abstractDownload PDF (730K) -
Naohisa KOCHIBE1973 Volume 74 Issue 6 Pages 1141-1149
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAn α-L-fucosidase [EC 3. 2. 1. -] has been purified from Bacillus fulminans by ammonium sulfate fractionation and subsequent column chromatography on Sephadex G-200, on DEAE-cellulose and on hydroxylapatite. Free fucose liberated by enzyme action was determined by microdiffusion method. The enzyme has a molecular weight of 70, 000 to 80, 000, hydrolyzes specifically a terminal, α-(1→2)-fucosidic bond of glycoproteins, glycopeptides and oligosaccharides and fails to cleave (1→3)- and (1→4)- linkages of α-L-fucosides in lacto-N-fucopentaose III and lacto-N-fucopentaose II, respectively, as well as p-nitrophenyl α-L-fucoside. It has apparent Km values of 3.3×10-4M, 1.6×10-3M, 2.0×10-3M, and 2.5×10-3M for bound fucose in desialyzed PSM, glycopeptide of desialyzed PSM, lacto-N-fucopentaose I and 2'-fucosyllactose, respectively.View full abstractDownload PDF (641K) -
Masataka YOSHINO, Toshio TAKAGI1973 Volume 74 Issue 6 Pages 1151-1156
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe molecular weights of native AMP nucleosidase [EC 3. 2. 2. 4] from Azotobader vinelandii and its subunits have been determined by sedimentation equilibrium analysis and gel-filtration. The molecular weight of the native enzyme, as determined by the first method, was 104, 000. The molecular weights of the subunits were determined by both methods in 6M guanidine hydrochloride (GuHCl)** containing dithiothreitol (DTT). The results obtained indicated that the enzyme is dissociated into subunits with identical molecular weights of 26, 000. It is therefore concluded that the native AMP nucleosidase molecule is composed of four polypeptide chains of essentially the same size. The active form of the enzyme is suggested to have an octameric structure taking into consideration of the results of our previous kinetic studies.View full abstractDownload PDF (420K) -
Yoshinori NOZAWA1973 Volume 74 Issue 6 Pages 1157-1163
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effects of a potent hypocholesteremic agent, p-chlorophenoxyisobutyrate (CPIB or clofibrate) on the growth, lipid content, and incorporation of 14C-labeled precursors (sodium [1-14C] acetate and [1-14C] palmitate) into various lipid fractions of Tetrahymena pyriformis, strain E, were investigated. Addition of CPIB induced a striking decrease in incorporation of 14C-labeled precursors into phospholipids. Synthesis from 14C-acetate of a sterol-like lipid, tetrahymanol, which is the most abundant neutral lipid and exclusively rich in the surface membranes, was also markedly inhibited by CPIB. Experiments with 14C-acetate showed increased incorporation of 14C-radioactivity into triglyceride associated with a corresponding decrease in phospholipid biosynthesis.View full abstractDownload PDF (485K) -
II. Electrophoretic, Kinetic, and Immunological Studies on Pyruvate Kinase of Erythrocytes and Other TissuesKiichi IMAMURA, Takehiko TANAKA, Toshihiro NISHINA, Koji NAKASHIMA, Sh ...1973 Volume 74 Issue 6 Pages 1165-1175
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Previous findings that three types of pyruvate kinase [EC 2. 7. 1. 40] (PK) are present in human tissues were confirmed and these PK's were partially characterized. Their distribution in human tissues was almost identical to that in rat tissues. Results of electrophoretic, immunological, and kinetic studies on erythrocyte PK suggested that it might be a hybrid of L and M2 subunits.
2. On the basis of these fundamental results, the PK in erythrocytes and other tissues (i.e., liver, muscle, spleen, and adipose tissues) of human subjects with two typical types of PK deficiency was analyzed. Case M.I. (PK “Beppu”) showed quantitative abnormality of erythrocyte PK and liver L-PK, while M1 (muscle) and M2 (spleen and adipose tissue) were normal. In cases Y. K. and F. K. (PK “Tokyo”), qualitative abnormality of erythrocyte PK and liver L-PK was found, while M1 (muscle) and M2 (spleen and adipose tissue) were normal as in case M.I.View full abstractDownload PDF (2975K) -
III. Ribonuclease H and Elongating Activity of Stimulatory Factor S-IIShunji NATORI, Kazuyuki TAKEUCHI, Den'ichi MIZUNO1973 Volume 74 Issue 6 Pages 1177-1182
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe characteristics of S-II, a factor stimulating partially purified RNA polymerase II, were studied further. It was found that S-II was inactivated by digestion with trypsin or heat-treatment at 60°C for 10min, indicating that it consists at least partly of protein.
No ribonuclease [EC 2. 7. 7. 16] or deoxyribonuclease [EC 3. 1. 4. 5] activity was detected in S-II, except ribonuclease H activity which was not separated from the stimulatory activity on a column of CM-cellulose. The ribonuclease H activity in the S-II preparation was repressed by MnCl2 without loss of stimulatory activity.
Using Ehrlich ascites tumor DNA as template it was found that the molecular size of RNA was much larger when synthesized in the presence of S-II. However, using calf thymus DNA as template the molecular size of the RNA synthesized was not affected by S-II.View full abstractDownload PDF (481K) -
Masasuke YOSHIDA, Tairo OSHIMA, Kazutomo IMAHORI1973 Volume 74 Issue 6 Pages 1183-1191
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSD-Fructose-1, 6-bisphosphatase [D-fructose-1, 6-bisphosphate 1-phosphohydrolase, EC 3. 1. 3. 11] from Flavobacterium thermophilum HB8, an extremely thermophilic bacterium, was partially purified. The preparation was free from non-specific phosphatase activity and its specific activity was 8 times that of the crude extract. The enzyme specifically hydrolyzed the C-1 phosphate ester bond of fructose 1, 6-bisphosphate to fructose 6-phosphate and inorganic phosphate. The enzyme was similar to the E. coli fructose 1, 6-bisphosphatase as regards optimum pH (8.1), requirement for divalentions, and Km for D-fructose 1, 6-bisphosphate (2×10-5M). The substrate, D-fructose 1, 6-bisphosphate, was inhibitory to the enzyme at high concentrations. The enzyme was activated by phosphoenolpyruvate, which released the substrate inhibition. AMP inhibited the enzyme activity, emphasizing the substrate inhibition. Inactivation by AMP was recovered to some extent by the addition of phosphoenolpyruvate. The enzyme was heat resistant and both the catalytic and allosteric properties of the enzyme remained unchanged after treatment at high temperatures, for instance, at 70°C for 1hr. The role of phosphoenolpyruvate in the regulation of glycolysis-gluconeogenesis in the bacterium is discussed.View full abstractDownload PDF (692K) -
Tomisaburo KAKUNO, Kazuo HOSOI, Tomihiko HIGUTI, Takekazu HORIO1973 Volume 74 Issue 6 Pages 1193-1203
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Cytochrome c2 in the soluble state transferred 1 electron (n=1) in the redox reaction at pH 5.5-9.8. Soluble and chromatophore-bound cytochrome cc' transferred 2 electrons at pH 5.0-5.5 and 1 electron at pH 7.0-9.5. The proton transfer number, (n-a), was different for soluble and bound cytochrome cc' over the pH range tested.
2. For bound ubiquinone-10, n was 2 over the pH range tested; the (n-a) values were 2 at pH 5.5-8.0 and 1 at pH 8.0-9.0.
3. The reduction of bound cytochrome cc' by succinate and the reduction of bound cytochrome B by NADH, were unchanged in D2O. However, the reduction of bound ubiquinone-10 by succinate and the reduction of added cytochrome c2 by NADH and by succinate were significantly inhibited.
4. The rates of ATP formation in the light, of ATP hydrolysis in the dark, and of ATP-Pi exchange in the dark, were inhibited to 41, 68, and 77%, respectively in 97% D2O.View full abstractDownload PDF (727K) -
Yukio NISIMOTO, Tomisaburo KAKUNO, Jinpei YAMASHITA, Takekazu HORIO1973 Volume 74 Issue 6 Pages 1205-1216
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIn Rhodospirillum rubrum chromatophores, oxidation of NADH by molecular oxygen was exhibited mostly by two different respiratory systems, one composed of NADH: hemeprotein oxidoreductase (enzyme-H), flavin, cytochrome B, and ubiquinone-10, and the other of NADH: quinone oxidoreductase (enzyme-Q) and flavin. These two systems were almost equal in their contribution to respiration. ATP formation was coupled with both systems. Added cytochrome c2 was reduced by the system involving enzyme-H, whereas 2, 6-dichlorophenol indophenol (DCPI) was reduced by the system involving enzyme-Q. Since the rate of oxidation was depressed to half when the chromatophores were incubated either with antiserum against enzyme-H (antiserum-H) or with antiserum against enzyme-Q (antiserum-Q), it seems probable that the enzyme-H and enzyme-Q molecules were bound on the surface of the chromatophore membrane in such a manner as to allow the antibodies to combine with their respective enzymes. The system involving enzyme-Q was able to reduce bound quinone at a relatively slow rate. In intact chromatophores, carbon monoxide did not combine with bound cytochrome B as well as bound cytochrome cc', although these hemeproteins were able to combine with carbon monoxide when the chromatophore membrane was impaired. Conceivably, the main sites responsible for the reduction of molecular oxygen were the quinone in the system involving enzyme-H, and the flavin in the system involving enzyme-Q.View full abstractDownload PDF (879K) -
I. The Key Role of Anions in Determining the Direction and Magnitude of Hydrogen Ion FluxYoshichika KOBAYASHI, Mitsuo NISHIMURA1973 Volume 74 Issue 6 Pages 1217-1226
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe mechanism of H+ transport in Rhodospirillum rubrum cells was investigated with respect to anion movement.
The cells of R. rubrum showed a light-induced H+ efflux, which required the intact cell membrane. The light-induced H+ efflux was observed only in the presence of appropriate anions, such as Cl- or N03-. The light-induced H+ efflux was inhibited by inhibitors of cyclic electron transport, antimycin A and o-phenanthroline, and uncouplers, carbonyl cyanide m-chlorophenylhydrazone (CCCP) and 2, 4-dinitrophenol (DNP) at higher concentrations.
The light-induced H+ efflux was stimulated by N-methylphenazonium methosulfate (PMS) and N, N, N', N'-tetramethyl-p-phenylenediamine (TMPD). PMS and TMPD partially overcame the inhibition by antimycin A and o-phenanthroline. The light-induced H+ efflux was stimulated by CCCP or DNP at lower concentrations in the presence of Cl- or NO3-.
When CCCP or DNP was added to the cell suspension in the dark, an uptake of H+ into the cells followed. The H+ uptake in the dark on the addition of CCCP or DNP also required the presence of Cl- or NO3-.
Direct measurement of 36Cl- translocation showed that Cl- is accumulated into the cells with H+ in the dark after the addition of CCCP.
These results suggest that the H+ movement couples with anion movement and that the transport of anions is the determining factor for the direction and magnitude of H+ flux.View full abstractDownload PDF (753K) -
II. Analysis of Reversed Hydrogen Ion ChangeYoshichika KOBAYASHI, Mitsuo NISHIMURA1973 Volume 74 Issue 6 Pages 1227-1232
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe direction of light-induced H+ transport in cells of Rhodospirillum rubrum was outward in NaCl solution. The direction of the light-induced H+ change was reversed by the addition of NH4+ and amine, and the reversal of H+ change was influenced by the pH of the medium. In NH4Cl solution, light-induced H+ transport was outwardly directed at acid pH (<pH 5.9), and inwardly directed at alkaline pH (>pH 5.9). At near pH 5.9 the light-induced H+ change showed a biphasic time-course. The two-directional H+ changes were inhibited by the addition of carbonyl cyanide m-chlorophenylhydrazone (CCCP) and antimycin A.
The pH value at which the direction of H+ change was reversed was dependent on the Cl-/NH4+ ratio in the medium. The reversal pH was 5.9 at Cl-/NH4+=1; it moved to the alkaline side on increasing the Cl-/NH4+ ratio.
Reversal of the H+ change was also observed in the presence of acetate or phosphate.
A possible mechanism of H+ transport in cells of Rhodospirillum rubrum was discussed in relation to anion and NH4+ transport.View full abstractDownload PDF (461K) -
III. The Influence of Uncouplers on Hydrogen Ion ChangeYoshichika KOBAYASHI, Mitsuo NISHIMURA1973 Volume 74 Issue 6 Pages 1233-1238
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWhen carbonyl cyanide m-chlorophenylhydrazone (CCCP) was added in the dark, a large amount of H+ was taken up into Rhodospirillum rubrum cells in the presence of appropriate anions (Cl- or N03-). The extent and the initial rate of the dark H+ influx depended on the bacteriochlorophyll and CCCP concentrations. Low concentrations of CCCP stimulated the light-induced H+ efflux of cells. The CCCP stimulation of the light-induced H+ efflux was highly pH dependent. At pH's higher than 7, the effect of CCCP was reversed and a slight inhibition of H+ efflux was observed.
Atebrin caused stimulation of light-induced H+ efflux of a type different from that of CCCP. Atebrin did not induce H+ changes in darkness.
A model for H+ transport in bacterial cells is presented and discussed with special reference to anion movement. The transport of anions is the determining factor for all types of H+ transport in the cell membrane of Rhodospirillum rubrum and Rhodopseudomonas spheroides.View full abstractDownload PDF (452K) -
Partial Purification and PropertiesTakashi SAIKI, Hideaki SHINSHI, Kei ARIMA1973 Volume 74 Issue 6 Pages 1239-1248
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Homoserine dehydrogenase [L-homoserine: NADP+ oxidoreductase] was extracted and partially purified 170-fold from an extreme thermophile, Thermus flavus AT-62.
2. The enzyme activity was not influenced by any single end product of the aspartate pathway nor by the simultaneous presence of these products, but was inhibited 85% by the presence of 10mM cysteine.
3. The enzyme was activated 4- to 5-fold by 500mM K+ or Na+, and the effect of K+ and Na+ was additive. Other monovalent cations, NH4+, Li+, Rb+, and Cs+ were less effective. Potassium or sodium ions also specifically protected the enzyme from heat inactivation. In the presence of 100mM K+, no loss of enzyme activity was observed after 30min at 70°C, and about 50% of the initial activity remained after 30min at 80°C.
4. The facts that the activation energy for the reaction below and above 50°C were 17, 000 and 8, 000cal per mole, respectively, and that negative homotropic cooperativity can be observed only at elevated temperatures (above 50°C), suggest that a temperature-dependent conformational change occurs in the enzyme protein at about 50°C.View full abstractDownload PDF (632K) -
VII. Characterization of the Reaction of Restoration of Active Transport Mediated by Binding ProteinYasuhiro ANRAKU, Hiroshi KOBAYASHI, Hiroshi AMANUMA, Akihito YAMAGUCHI1973 Volume 74 Issue 6 Pages 1249-1261
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSStudies were made on the protein factors released by osmotic shock, which were active in restoration of active transport in osmotically shocked cells of Escherichia coli.
Binding proteins were essential for the restoration reaction. Leucine-binding protein-I, which binds leucine, isoleucine, valine, and threonine restored active uptake of leucine and isoleucine in shocked cells. Leucine-binding protein-II, which binds leucine, was active for restoration of leucine uptake, but not for that of isoleucine. 3H-Acetylated leucine-binding protein-I was prepared and found to have the restorative activity.
The restorative activity of the galactose-binding protein was examined using various mutants in which the transport carrier systems have been characterized. This binding protein restored galactose uptake, but not glucose uptake in W3092 cells. It restored both galactose and glucose uptake in W2243A cells, but not in S7 cells. All these results are consistent with the concept that binding proteins are functional components of these transport carrier systems.
Two restorative factors other than binding proteins were also separated by gel filtration from the shock fluid. One of them was associated with a component of high molecular weight containing adenosinetriphosphatase activity. It was concluded that membrane-bound adenosinetriphosphatase [EC 3. 6. 1. 3] is involved in the reaction for restoration of leucine uptake. The other factor was identified as a non-dialyzable protein of low molecular weight. It was only active in the restoration reaction when added with phospholipid of E. coli.
The effects of various compounds, including the carbon source, inorganic ions, nucleotides, and polyamines on the restoration reaction were examined.View full abstractDownload PDF (1666K) -
Mitsuo NAKAGAWA, Toshiro NISHIDA1973 Volume 74 Issue 6 Pages 1263-1266
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
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Haruko FUJIMORI, Keitaro HIROMI, Shunsaku KOYAMA, Yoshihide MOMOTANI, ...1973 Volume 74 Issue 6 Pages 1267-1270
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (1076K) -
Masatake OHNISHI, Toshihiko SUGANUMA, Haruko FUJIMORI, Keitaro HIROMI1973 Volume 74 Issue 6 Pages 1271-1273
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (221K) -
Kazuo HOSOI, Setsuko YOSHIMURA, Gilbu SOE, Tomisaburo KAKUNO, Takekazu ...1973 Volume 74 Issue 6 Pages 1275-1278
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (266K) -
Kenji TAKAHASHI1973 Volume 74 Issue 6 Pages 1279-1282
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (282K) -
Tsuneo KAMEYAMA, Takamitsu SEKINE1973 Volume 74 Issue 6 Pages 1283-1285
Published: December 25, 1973
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (230K)
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