1. A method was described for the purification of ATP deaminase [ATP aminohydrolase, EC class 3. 5.4] from wheat-bran koji of Microsporum audouini.
2. This enzyme was found to deaminate adenosine tetraphosphate, ADP, 5'-AMP, ADP-ribose, ADP-glucose, dATP, dADP, dAMP, 3', 5'-cyclic AMP, NAD, FAD, CoA, and adenosine, as well as ATP. Adenine, 2'-AMP, and NADP were not attacked by this enzyme.
3. The deaminated product of ATP was proved to be ITP.
4. The stoichiometry and equilibrium of deamination were studied.
5. The activation energy of the enzyme activity was found to be approximately 7, 300 calories.
6. The optimal pH was 5.0 in acetate buffer.
7. The Michaelis-Menten constant was calculated to be 4×l0^-5M at 30°C.

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A modified phosphotransacetylase method for the determination of CoA using phosphotransacetylase [EC 2. 3. 1. 8] from Escherichia coli B is described.
This method has a high specificity for CoA.
Diluted solutions of phosphotransacetylase from this source are inactivated during storage, but the enzyme is stable in the presence of sulfate ion.
The inhibitory effects of sulfate ion and other inorganic salts on the enzyme activity are also reported.

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The mechanism of action of mikamycins A and B, inhibitors of protein synthesis, was investigated with E. coli ribosomal systems. In both poly U- and poly A- systems polypeptide synthesis by ribosomes charged with the respective polyribonucleotides was as sensitive to mikamycins A and B as that by ribosomes which had been preincu-bated with the antibiotics prior to the addition of polyribonucleotides. This fact suggests that both kinds of mikamycin may not affect the formation of the ribosome-polyribonucleotide complex.
Mikamycin A inhibited the binding of several C¹⁴-aminoacyl-sRNA's tested to the ribosome-polyribonucleotide complex to varied extents. The binding of phenylalanyl- and lysyl-sRNA's was more markedly affected than that of prolyl-sRNA and the extent of such inhibitions of binding among these three systems was parallel to, though less than, that of the corresponding over-all polypeptide synthesis. On the contrary, mikamycin B did not significantly inhibit the attachment of any aminoacyl-sRNA tested to the active complex. The degree of inhibition by mikamycin A of polypeptide synthesis decreased when it was introduced to the reaction mixture later, whereas that by mikamycin B showed a tendency to increase with the delay of addition. These results suggest that mikamycin A primarily affects an early ribosomal phase of chain growth involving the binding of aminoacyl-sRNA and the initial peptide chain growth, whereas the site of action of mikamycin B is thought to be in somewhere after the phase attacked by mikamycin A.

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1. Liver microsomes were obtained from both phenobarbital-treated and untreated rabbits, and used for the studies on the spectral properties of P450.
2. The absorption spectra of the control and the induced microsomes were measured using, as the reference of spectrophotometry, the microsomes which were decolorized by the hydrogen peroxide-treatment.
3. The spectrum of the induced components was obtained by taking the difference spectrum, induced minus control, with the preparations which were so adjusted as to contain the same amount of cytochrome b₅.
4. The approximate absorption spectrum of P-450 in liver microsomes was obtained photometrically by subtracting the spectrum of the preparation, which had been subjected to a prolonged aerobic treatment with pCMB, from the spectrum of the untreated control. The resulting spectrum showed essentially the same characteristics as that of P-450 in adrenal cortex mitochondria.
5. The nature of the colored prosthetic group of P-450 and the limitation in the accuracy of measurement performed in the present study were discussed.

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1. Conjugation of myosin A with 1-dimethylaminonaphthalene-5-sulfonyl chloride (DNS) produced a quenching of the tryptophan fluorescence depending on the degree of labeling. It was shown that the degrees of quenching of the ultraviolet fluorescence by labeling with the dye in different proteins were almost of the same order of magnitude if the degree of labeling is expressed as “moles dye per mole tryptophan”.
2. The ultraviolet fluorescence of DNS-labeled myosin A showed a significant quenching in the pH region in which “normal” tyrosine ionize. The quenching of the visible fluorescence excited by the irradiation in the tryptophan region (285mμ) approximately paralleled the ionization zone of “abnormal” tyrosine. The DNS fluorescence excited at 330mμ was slightly enhanced in these alkaline pH region. In the presence of ethanol, these alkaline quenching curves were shifted slightly to lower pH values.
Urea decreased the ultraviolet and the visible fluorescence at pH's below about 10. However, it reduced the alkaline quenching of both fluorescence intensities and enhanced the visible fluorescence excited at 330mμ in this alkaline pH region.
The visible fluorescence intensity excited at 330mμ decreased more extensively than that excited at 285mμ at pH 6.
3. The time-dependence of the fluorescence intensities was studied after the addition of dioxane, ethanol, and urea. Furthermore, the effects of urea and guanidine hydrochloride on the intensities were measured at different concentrations of the reagents.
The results obtained showed that the measurements of the fluorescence intensities of both the tryptophanyl residues of myosin A and DNS attached to the enzyme provides a useful means to monitor the structural transitions of the protein.

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1. RNase T₂ [EC 2. 7. 7. 17] has no absolute specificity as regards nucleotide bases, although it shows a preference for adenylic acid linkages in RNA.
2. On exhaustive digestion with RNase T₂, ribonucleic acids and homopolynucleotides are quantitatively digested to produce nucleoside 3'-phosphate.
3. In the partial digestion of RNA with RNase T₂, some phosphodiester bonds of Ap in RNA are preferentially split to produce the large fragments terminating with Ap in the very early stage of digestion, while few of the other nucleotide bonds are split.
4. The susceptibility to RNase T₂ is different depending on the molecular conformation of the substrates used.

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The primary structures of the cytochromes c from Saccharomyces oviformis, S. cerevisiae and their respiration-deficient mutants were compared by the criteria of the chromatograms on Amberlite CG-50 columns, amino acid compositions and peptide maps of the tryptic digests. It was concluded that these strains contained a cytochrome c of the same primary structure.

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1. By a combination of acetone fractionation, ammonium sulfate fractionation and chromatography on DEAE-cellulose, bacterial phosphopyruvate hydratase [EC 4. 2. 1. 11, enolase] isolated from Br. fuscum was purified to about 32.3-fold the activity of the cell-free extract.
2. The purified enzyme is more unstable than that of yeast or muscle, and rapid loss of activity was observed on heating at 50°C for 5 minutes.
3. The metal ion specificity of bacterial phosphopyruvate hydratase was investigated and the following order was found for the effectiveness of various metals:
Mg⁺⁺>Zn⁺⁺>Fe⁺⁺>Mn⁺⁺>Cu⁺⁺>Ni⁺⁺>Ag⁺>Co⁺⁺>Cd⁺⁺
In addition to the ions already known to activate the yeast enzyme, Cu⁺⁺ and Ag⁺ were found to be effective for activation of bacterial phosphopyruvate hydratase.
4. Other metals tested, except Hg+, were found to inhibit Mg⁺⁺-phosphopyruvate hydratase, their effects being in the following order:
Fe⁺⁺⁺>Hg⁺⁺>Zn⁺⁺>Co⁺⁺>Fe⁺⁺>Ca⁺⁺>Mn⁺⁺>Cu⁺⁺>Al⁺⁺⁺>Ag⁺>
Ni⁺>Mo^vi>Pb⁺⁺>Ba⁺⁺
However, it is striking that only Hg+ could increase the activity of Mg⁺⁺-phosphopyruvate hydratase.
5. The optimum pH for the rate of PEP turnover was found to be 6.9-7.0.
6. The Michaelis constant for PEP was 4.2×10^-4M, and the concentrations of PEP above 1×10^-2M inhibited the enzyme.
7. The Michaelis constants for the reverse reaction were 4.8×10^-4M for Mg⁺⁺ and 7.9×10^-5M for Zn⁺⁺. At a Mg⁺⁺ concentration above 2.56×10^-2M, no inhibition could be observed, while Zn⁺⁺ concentrations higher than 1.1×10^-3M were inhibitory.
8. In the presence of both Mg⁺⁺ and Zn⁺⁺ a more complicated inhibition, which was neither simply competitive nor non-competitive inhibition, was observed.
9. The apparent activation energy of the reverse reaction of Mg⁺⁺-phosphopyruvate hydratase (PEP→2-PGA) was calculated to be 10, 680 cal./mole. This value is considerably smaller than that obtained for the forward reaction of the yeast enzyme.
10. The bacterial phosphopyruvate hydratase was found to be inhibited by several kinds of chemicals such as sodium floride, phloroglucinol, 8-hydroxyquinoline, EDTA, phioridine and reduced glutathione, while PCMB and monoiodoacetate had no effect on the activity.

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1. A ribonuclease [EC 2. 7. 7. 17] involved in some unknown manner in the debris fraction of Escherichia coli strain B was purified 12, 500 fold by treatment with 4M urea and 0.05M NaCl, ammonium sulfate fractionation at pH 4.5 and chromatography on CG-50 resin. Its properties and mode of action were studied in comparison with those of latent ribosomal ribonuclease purified 700 fold by the same procedures.
2. The ribonuclease from the debris was adsorbed on a CG-50 column and eluted with 0.2M potassium phosphate buffer, pH 7.4, containing 0.4M sodium chloride. It was found that the enzyme has the same catalytic action as ribosomal ribonuclease. However, distinct differences between debris and ribosomal ribonuclease were found in their selectivity to hydrolyze substrate RNA's. The debris ribonuclease acted preferentially on small molecules of ribonucleic acid, while the ribosomal ribonuclease preferentially hydrolyzed ribonucleic acid molecules of high molecular weight.
3. The debris ribonuclease was more heat-stable than the ribosomal ribonuclease. However, both enzymes were inhibited by urea and activated by sodium chloride and they had the same optimal pH range of 7.4 to 7.8.
4. It was suggested that the debris ribonuclease may form a lysozyme sensitive complex with components of the cell membrane. The possible mechanism of location of ribonuclease is discussed on the basis of the results.

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1. The relationship between ribonuclease [EC 2. 7. 7. 17, E. coli] and cyclic phosphodiesterase [EC class 3. 1. 4] in the metabolism of ribonucleoside 2', 3'-cyclic phosphate was studied. Ribonuclease and cyclic phosphodiesterase hydrolyze, respectively, adenosine 2', 3'-cyclic phosphate at rates of 0.01 and 11.5μmoles per hour per unit of enzyme. Using these values, the capacity of the cells to hydrolyze ribonucleoside 2', 3'-cyclic phosphate was estimated and it was found that cyclic phosphodiesterase was the chief enzyme which catabolizes the above cyclic nucleotides.
2. Cyclic phosphodiesterase was found to act on oligonucleotides with cyclic phosphate at their 2', 3'-hydroxyl ends. Hydrolysis of their cyclic phosphate to the corresponding 3'-isomers was a rapid process compared with the subsequent dephosphorylation. The longer the chain length, the slower was the reaction velocity of dephosphorylation.
3. The mechanism of adsorption of ribonuclease onto the ribosomes was studied. The ribonucleases from the debris and ribosomes were adsorbed similarly and the ribosomes could adsorb up to about 10 times as much enzyme as was initially present on them. Bovine pancreatic ribonuclease A [EC 2.7.7.16] and ribonuclease T1 [EC 2.7.7.26] were not adsorbed well by the ribosomes of E. coli.
4. Based on the results it is proposed that a ribonuclease-cyclic phosphodiesterase system is one enzymatic pathway for degradation of ribonucleic acid in vivo. A possible mechanism regulating this system is discussed.

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1. Using highly purified phospholipid preparations, the effects of various salts on the surface pressure and the surface potential of the phospholipid monolayer are studied. The results are in good agreement with those obtained by several authors.
2. The surface potential of cardiolipin decreases at high compression region in the presence of acetylcholine.
3. The results of preliminary experiments on the action of acetylcholine show that the decrease of the surface potential of cardiolipin monolayer is enhanced by Ca, but suppressed by Na or hexamethonium bromide.

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1. In growing cells of Mycobacterium phlei, phospholipids showed an appreciable turnover.
2. Among the components of phospholipids, cardiolipin had the highest turnover rate.
3. Phosphatidyl-inositol oligomannosides and phosphatidyl ethanolamine showed relatively low turnovers compared with that of cardiolipin.
4. Phosphatidic acid was also a metabolic precursor of phospholipids even in the cells of M. phlei.

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1. The concentrations of glycolytic intermediates and adenine nucleotides in reticulocytes from rabbits which were induced by acetylphenylhydrazine injection, were analyzed and compared with those of the mature cells. The following changes were found to be correlated with the reticulocyte count of blood, when the samples were collected during the in vivo maturation of the cells: The concentrations of hexose-6-phosphates in reticulocytes are high compared with those of the mature cells, whereas those of fructose diphosphate, triose phosphates, monophosphoglycerates and phosphoenolpyruvate show little difference in reticulocytes and mature red cells. The concentration of α-glycerophos-phate is found to be extremely high compared with that in the mature cells. The content of ATP is high in reticulocytes, while that of 2, 3-diphosphoglycerate is lower than that in the mature cells.
2. The effect of acetylphenylhydrazine to mature rabbit erythrocytes incubated with glucose was studied to exclude the possibility that the above observations were caused by the reagent. The level of ATP as well as those of the hexose monophosphates decreased on the addition of acetylphenylhydrazine, while that of 3-phosphoglycerate increased.
3. The lactate formation as well as the pattern of glycolytic intermediates and nucleotides in reticulocytes incubated in the presence of glucose and inorganic phosphate at pH 7.4 were analyzed. On the addition of cyanide, the concentrations of hexose 6-phosphates in the cells decrease and at the same time the lactate formation increases. This agrees with the observations made with other kind of cells. The addition of 2, 4-dinitrophenol also causes an increase of the lactate formation with the concomitant decrease of the glycolytic intermediates including hexose 6-phosphates.

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It has previously been shown that deoxy-ATP can induce contraction of myofibrils and the enzymatic activity of actomyosin type, and that deoxy-ATP is scarcely incorporated into G-actin. In this report, the superprecipitation of actomyosin by deoxy-ATP was shown to be sensitive to EGTA. The time-course of P1-liberation from the myosindeoxy-ATP system showed an initial burst of about I mole per 4×10⁵g. of myosin which originated from the decomposition of a reactive myosinphosphate complex caused by trichloroacetic acid. Therefore, to clarify the role of ADP bound to F-actin in contraction of muscle models, we investigated the superprecipitation and the enzymatic activity of reconstituted actomyosin made with myosin and ADP free F-actin, which was prepared by the method of Kasai et al. or of Mommaerts, using deoxy-ATP as the substrate.
When Kasai's F-actin free from ADP was used, the deoxy-ATP induced superprecipitation of actomyosin was scarcely affected, but the enhancement of deoxy-ATPase activity of myosin by F-actin was decreased remarkably by the decrease in the ADP content of F-actin. When F-actin completely free from ADP was used, only 0.1 mole of deoxy-ATP per mole of myosin was hydrolyzed by the actomyosin type deoxy-ATPase, before the superprecipitation of actomyosin was completed. From these results, we concluded that the enzymatic activity of actomyosin type is unnecessary for the superprecipitation. Measurements of the light-scattering of the myosin-F-actin system showed that Kasai's F-actin free from ADP scarcely binds to myosin.
Not only the superprecipitation induced by deoxy-ATP but also the deoxy-ATPase activity of actomyosin reconstituted from myosin and Mommaerts' F-actin free from ADP were almost the same as those of the control actomyosin. Furthermore, this type of ADP free F-actin could bind to myosin to almost the same extent as the control F-actin. Therefore, it was concluded that the disappearance of actomyosin type deoxy-ATPase activity of actomyosin made with myosin and Kasai's F-actin free from ADP was not directly due to the removal of ADP but to a conformational change in actin occurring during the removal of nucleotide from G-actin by the method of Kasai et al.
When ATP was used as the substrate, the ATPase activity of myosin was considerably enhanced even by Kasai's F-actin free from ADP, and no significant difference in the enzymatic activities of the control actomyosin. and actomyosin free from ADP was observed, if the ratio of F-actin to myosin was higher than 1.5:8. This result suggests that ATP or ADP was incorporated into Kasai's F-actin during the superprecipitation of actomyosin.

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The dependence of superprecipitation and ATPase activity of myosin B on the concentration of ATP was studied in the presence and absence of pyruvate kinase and phosphoenolpyruvate. The minimum concentration of ATP required to maintain the maximum extent of superprecipitation of myosin B decreased on addition of the kinase system. The rate of superprecipitation was proportional to 1.7th power of the ATP concentration. Both the rate of superprecipitation and the ATPase activity increased with increase in the concentration of free Ca⁺⁺, but were unaffected by free EGTA. ADP and P₁ accelerated the rate of superprecipitation of myosin B without enhancing the ATPase activity. In the presence of a sufficient amount of ADP and P₁, the extent of superprecipitation was proportional to the concentration of myosin B, but the specific rate of superprecipitation decreased with decrease in the myosin B concentration and superprecipitation did not occur at protein concentrations lower than a critical one.
The rate of superprecipitation of a large amount of myosin B was conspicuously accelerated by the addition of an extremely small amount of previously superprecipitated myosin B. The accelerating effect of superprecipitated myosin B was markedly increased by sonication. The rate of superprecipitation of 0.3mg. per ml. of myosin B induced by ATP was increased 20-30 fold by the addition of 0.3μg. per ml. of superprecipitated and sonicated myosin B. Neither the precipitate dissolved in 0.6M KCl nor the supernatant of superprecipitated myosin B after centrifugation accelerated the rate of superprecipitation. A minute amount of superprecipitated and sonicated myosin B added as an accelerator of the superprecipitation had no effect on the ATPase activity. In the presence of a minute amount of superprecipitated and sonicated myosin B, the specific rate of superprecipitation became independent of the concentration of myosin B but it was proportional to the concentration of the accelerator up to a critical concentration.
On the basis of these results, it was concluded that superprecipitation is a nucleated growth process and a minute amount of previously superprecipitated myosin B serves as a nucleus for the superprecipitation of a large amount of myosin B.

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1. A new bile acid, haemulcholic acid was isolated from the bile of Parapristipoma trilineatum.
2. The acid was revealed to have the chemical constitution of 3α, 7α, 22β-trihydroxy-5β-cholanic acid.

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