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Magnetic domain states of completely isolated particles of barium ferrite between 0.1 and 2 μm in size are investigated by electron holography. Phase distribution is reconstructed digitally by the Fourier transform method and converted to interference micrographs. In the interference micrographs single magnetic‐domain particles are observed directly and clearly for the first time. Furthermore, it is found that single‐domain and two‐domain particles coexist down to sizes as small as about 0.1 μm.

Among the yeasts belonging to the genus Torulaspora, T. pretoriensis IFO 10218 was selected as a maltose-fermenting strain that could leaven dough containing 5% glucose as high as commercial baking strains. The freeze-thaw resistance of this strain in the dough was higher than that of Saccharomyces cerevisiae FL 2209 isolated from commercial compressed yeast. Since the cells of IFO 10218 readily sedimented in their suspension, a dispersing mutant, YK-1, was induced. When YK-1 was cultured in the. presence of NaCl, leavening abilities were reduced, unlike FL 2209. α-Glucosidase activity of YK-1 cells grown on maltose increased slightly, resulting in partly improved leavening ability in the dough without addition of sugar. YK-1 was shown to be a candidate applicable to breadmaking by not only usual method but also the frozen-dough method.

(1992). Purification, Crystallization, and Properties of the Extracellular Levansucrase from Zymomonas mobilis. Bioscience, Biotechnology, and Biochemistry: Vol. 56, No. 8, pp. 1335-1337.

Rat cytochrome P -4501A1-dependent monooxygenase activities were examined in detail using recombinant yeast microsomes containing rat cytochrome P -4501A1 and yeast NADPH- P -450 reductase. On 7-ethoxycoumarin, which is one of the most popular substrates of P -4501A1, the relationship between the initial velocity ( v ) and the substrate concentration ([S]) exhibited non-linear Michaelis-Menten kinetics. Hanes-Woolf plots ([S]/ v vs. [S]) clearly showed a biphasic kinetic behavior. Aminopyrine N -demethylation also showed a biphasic kinetics. The regression analyses on the basis of the two-substrate binding model proposed by Korzekwa et al. (Biochemistry 37 (1998) 4137–4147) strongly suggest the presence of the two substrate-binding sites in P -4501A1 molecules for those substrates. An Arrhenius plot with high 7-ethoxycoumarin concentration showed a breakpoint at around 28°C probably due to the change of the rate-limiting step of P -4501A1-dependent 7-ethoxycoumarin O -deethylation. However, the addition of 30% glycerol to the reaction mixture prevented observation of the breakpoint. The methanol used as a solvent of 7-ethoxycoumarin was found to be a non-competitive inhibitor. Based on the inhibition kinetics, the real V max value in the absence of methanol was calculated. These results strongly suggest that the recombinant yeast microsomal membrane containing a single P -450 isoform and yeast NADPH- P -450 reductase is quite useful for kinetic studies on P -450-dependent monooxygenation including an exact evaluation of inhibitory effects of organic solvents.

A phase‐shifting method to directly measure the amplitude and phase distribution of specimen in an electron holographic microscope without the introduction of large number of carrier fringes is proposed. The initial phase of electron holograms is shifted by tilting the incident electron beam with a digital voltage/current supply. Several holograms of a specimen with properly different phase shifts are recorded digitally and used to calculate the amplitude and phase distributions of the specimen. Experimental result of observing the phase distribution of a cubic MgO crystal is shown.

Porcine pancreatic alpha-amylase (1,4-alpha-D-glucan glucanohydrolase) [EC 3.2.1.1] has both amylase activity (hydrolysis of alpha-1,4-D-glucoside bond of starch) and maltosidase activity (hydrolysis of p-nitrophenyl-alpha-D-maltoside to p-nitrophenol and maltose). By the modification of histidine residues of porcine pancreatic alpha-amylase with diethylpyrocarbonate (DEP), both amylase and maltosidase activities were decreased in the absence of chloride ion. In the presence of chloride ion, however, maltosidase activity of the modified enzyme was increased to more than 260% of that of the native enzyme, whereas amylase activity was decreased to less than 15% of the native enzyme. Since the chloride ion binding site is part of the active site loop [Buisson et al. (1987) Food Hydrocolloids 1,399-406 and Buisson et al. (1987) EMBO J. 6, 3909-3916], the special arrangements of both catalytic and modified histidine residues induced by the chloride ion binding would enhance only the maltosidase activity of the histidine-modified enzyme.

Fluoride-ion-promoted hydrolysis of phosphoric acid esters and amides to give phosphorofluoridic and phosphoramide fluoridic acid monoester salts takes place upon treatment with a THF solution of T...

Thermolysin is remarkably activated in the presence of high concentrations (1-5 M) of neutral salts [Inouye, K. (1992) J. Biochem. 112, 335-340]. The activity is enhanced 13-15 times with 4 M NaCl at pH 7.0 and 25 degrees C. In this study, the effect of neutral salts on the solubility of thermolysin has been examined. Although the solubility was only 1.0-1.2 mg/ml in 40 mM Tris-HCl buffer, pH 7.5, in the temperature range between 0 and 60 degrees C, it was increased greatly by the addition of salts. With NaCl, the solubility showed a bell-shaped behavior with increasing NaCl concentration, and the maximum solubility (10 mg/ml) was at 2.0-2.5 M NaCl. With LiCl and NaI, it increased progressively to 20-50 mg/ml with increasing salt concentration up to 5 M. The solubility observed in the presence of salts decreased with increasing temperature from 0 to 60 degrees C, and also with the order of chaotropic anion effect. The molecular weight of thermolysin was estimated to be 33.0(+/-2.5) x 10(3) in the presence of 0-3 M NaCl, suggesting that thermolysin exists as a monomer in the presence or absence of 3 M NaCl. The possibility that aggregation and/or dispersion of thermolysin might be related to the remarkable activation by salt was ruled out.

Poly(3-hydroxybutyrate) depolymerase was purified to homogeneity from the culture filtrate of Paecilomyces lilacinus D218 by column chromatography on CM-Toyopearl 650M and hydroxylapatite. The molecular weight of the enzyme was estimated to be 48,000 by SDS-PAGE. Maximal activity was observed near pH 7.0 and 45°C. The K m and V max values for PHB were 0.13 (mg/ml) and 3750 (U/mg protein), respectively. The enzyme hydrolyzed PHB and p-nitrophenyl fatty acids but not polycaprolactone and triglycerides.

The states of 28 tyrosyl residues of thermolysin have been characterized by means of pH-jump studies and nitration with tetranitromethane. The ionization states of phenolic groups of the tyrosyl residues have also been estimated by spectrophotometric titration of the absorption change at 295 nm. The ionization of 16 tyrosyl residues was completed within 15 s after a pH-jump, and these residues are considered to be located on the surface of thermolysin. On the other hand, the ionization of the other 12 residues required 15 s to 10 min, suggesting the occurrence of a conformational change which leads to exposure of the buried tyrosyl residues to the solvent. Sixteen tyrosyl residues were nitrated and categorized into three classes according to reactivity. The second-order rate constants of the respective classes of tyrosyl residues for nitration were evaluated as 3.32, 0.52, and 0.18 M-1.min-1, and their apparent pKa values were estimated to be 10.2, 11.4, and 11.8. Tyrosyl residues in the first class were considered to be located almost freely on the surface, while those in the second and third classes might be in constrained states.

The activity of thermolysin in the hydrolysis of N-[3-(2-furyl)acryloyl]-glycyl-L-leucine amide and N-carbobenzoxy-L-aspartyl-L-phenylalanine methyl ester is remarkably enhanced in the presence of high concentrations (1–5 M) of neutral salts [Inouye (1992) J. Biochem. (Tokyo) 112, 335–340]. In this study, the effect of salts on such activity has been examined using a series of substrates, furylacryloyl dipeptide amides, which have various hydrophobic amino acids at the cleavable bond. Although the enzyme activity varies widely depending on the substrate employed, the degree of activation at a given concentration of NaCl is considerably similar. This indicates that the degree of activation is not dependent on the hydrophobicity of the amino acid side chains at the scissile bond of the substrates. The molecular activity, kcat, and Michaelis constant, Km, were evaluated separately for substrates N-[3-(2-furyl)acryloyl]-L-leucyl-L-alanine amide and N-[3-(2-furyl)acryloyl]-L-phenylalanyl-L-alanine amide, and the activation was found to be brought about only by an increase in kcat. The effectiveness of monovalent cations on the increase of kcat was determined to follow the order of Na+ > K+ > Li+.

Lysyl-tRNA synthetase [L-lysine:tRNA(Lys)ligase (AMP forming); EC 6.1.1.6] was purified from Bacillus stearothermophilus NCA1503 approximately 1,100-fold to homogeneity in PAGE. The enzyme is a homodimer of M(r) 57,700 x 2. The molar absorption coefficient, epsilon, at 280 nm is 71,600 M-1.cm-1 at pH8.0. Enzyme activity in the tRNA aminoacylation reaction and the ATP-PPi exchange reaction increases up to 50 degrees C at pH 8.0, but is lost completely at 70 degrees C. The pH-optima of the two reactions are 8.3 at 37 degrees C. In the tRNA aminoacylation reaction, the Km values for L-lysine and ATP are 16.4 and 23.2 muM, respectively, and in the ATP-PPi exchange reaction, the Km values for L-lysine and ATP are 23.6 and 65.1 muM, respectively at 37 degrees C, pH 8.0. Interaction of either L-lysine or ATP with the enzyme has been investigated by using as a probe the ligand-induced quenching of protein fluorescence and by equilibrium dialysis. These static analyses, as well as the kinetic analysis of the L-lysine dependent ATP-PPi exchange reaction indicate that the binding mode of L-lysine and ATP to the enzyme is sequential ordered (L-lysine first). The interaction of lysine analogues with the enzyme has also been investigated.

A binuclear copper(II) phthalocyanine complex, in which two Cu(II) phthalocyanine units are linked by their common benzene ring to give a planar structure, was prepared and investigated concerning ...

Thirteen kinds of metal oxides were tested for one-pot selective synthesis of esters from nitriles and alcohols. Ceria (CeO2) showed more than two orders of magnitude higher activity than the other oxides. CeO2 acted as a reusable and effective catalyst for the ester synthesis from various nitriles and alcohols under neutral and solvent-free conditions at 160 °C. This method provides a rare example for the synthesis of heteroaromatic esters, which have been difficult to synthesize by conventional catalytic esterification methods. Valuable esters such as picolinic acid alkyl esters and niacin benzyl esters were synthesized, demonstrating a practical aspect of the present method. Kinetic studies suggested the following reaction mechanism: (1) H2O and ROH dissociate on CeO2, (2) nucleophilic attack of hydroxyl species (OHδ−) to the adsorbed nitrile on CeO2, leading to the formation of the primary amide, (3) nucleophilic attack of alkoxide species (ORδ−) to the amide as the rate-limiting step.

Among various metal oxides, cerium oxide (CeO2) shows the highest catalytic activity for transamidation of picolinamide with n-octylamine. CeO2 acts as a reusable and effective heterogeneous catalyst for transamidation under solvent-free conditions. Transamidation of a variety of amides and amines produced the corresponding N-alkyl amides in high yields. This method provides the first example of a heterogeneous catalyst for transamidation using aliphatic amines as substrates. Characterization of acid–base properties and kinetic studies suggest that the cooperation of the weak Lewis acid sites and adjacent strong base sites play important roles in the transamidation reaction.

Magnetic lines of force penetrating a superconducting thin foil have been investigated by means of electron holography. A field-emission TEM with a specially constructed cold stage was used to cool a Nb thin foil down to 4.5 K and apply magnetic fields up to 100 G. The specimen is tilted by 45° to both the electron beam and the magnetic field (applied horizontally) allowing the 2-D lattice of penetrating flux-lines to be discerned. The phase distribution of electrons transmitted through the specimen were quantitatively measured. Interference micrographs revealed tiny regions where the phase distribution rapidly changed. These regions coincided spatially with the spot-like contrast observed by Lorentz microscopy and were found to be quantized vortices containing a flux of h/2e. The experimental results were in good agreement with those predicted by theoretical simulations. Experiments exploring the vortex inner core structure at high resolution are presented.

Magnetic domain structures of barium ferrite particles are studied by electron holography. Single magnetic-domain particles have been observed directly and clearly for the first time. Furthermore, an on-line real-time electron holography system has been developed for dynamic observation. Using the system, magnetic domain-walls in a thin permalloy film have been observed in real-time. Dynamic behavior of magnetic domain-walls in the magnetization or demagnetization processes is clearly observed.

An Al2O3/SiO buffer layer was incorporated in a metal–insulator–semiconductor (MIS) Ta2O5 capacitor for dynamic random access memory (DRAM) application. Al2O3 was chosen for the buffer layer owing to its high band offset against silicon and oxidation resistance against increase in effective oxide thickness (EOT). It was clarified that post-deposition annealing in nitrogen at 800 °C for 600 s increased the band offset between Al2O3 and the lower electrode and decreased leakage current by two orders of magnitude at 1 V. Furthermore, we predicted and experimentally confirmed that there was an optimized value of y in (Si3N4) y (SiO2)(1-y), which is 0.58, for minimizing the leakage current and EOT of SiON. To clarify the oxidation resistance and appropriate thickness of Al2O3, a TiN/Ta2O5/Al2O3/SiON/polycrystalline-silicon capacitor was fabricated. It was confirmed that the lower electrode was not oxidized during the crystallization annealing of Ta2O5. By setting the Al2O3 thickness to 3.4 nm, the leakage current is lowered below the required value with an EOT of 3.6 nm.

Lysine residues of porcine pancreatic alpha-amylase (PPA) were modified with trinitrobenzenesulfonate (TNBS). 6 out of 21 lysine residues were modified per PPA molecule. Amylase activity (hydrolysis of the alpha-1,4-D-glucoside bond) was decreased to about 50% of the native enzyme, as judged from the kcat value at pH 6.9 after the modification, whereas maltosidase activity (hydrolysis of p-nitrophenyl-alpha-D-maltoside producing p-nitrophenol and maltose) was increased to about 250%. The increase in maltosidase activity of the modified PPA was due to the increase in kcat, but not to the decrease in Km. Modification of PPA with five kinds of acid anhydrides also caused the same effect as TNBS, including the number of modified lysine residues. The degree of increase in maltosidase activity was fairly proportional to the volume of the incorporated modification reagent. A modification protection study in the presence of maltotriitol (G3OH), which protected two out of six modifiable lysine residues against modification, suggested that a lysine residue at the substrate-binding site contributes to the change of substrate specificity.

The basic principles and methods of off-axis electron holography are presented and illustrated by means of three examples related to its application in high resolution electron microscopy and the investigation of electric and magnetic fields in thin specimens.

The micromagnetic structure and magnetization process of perpendicular Y3Fe5O12 (YIG) films were studied by Lorentz microscopy and electron holography. The closure domain structure inside the thin transmission electron microscopy specimen exhibits the same period as the magnetization pattern observed by magnetic force microscopy indicating the perpendicular anisotropy of the YIG film. Through observation of stray fields, it is concluded that the shapes of domain and domain walls are sensitive to the specimen thickness; moreover, a closure domain configuration observed in thin specimen is the stable energy state as determined by the balance between the crystalline anisotropy and shape anisotropy. Domain wall movement is observed by applying a magnetic field, in situ, inside the microscope in both horizontal and perpendicular directions; the saturation fields observed are qualitatively in agreement with the results of the hysteresis loop.

An imaging technique for reconstructing the projected potential of crystals using high-resolution electron holography is developed. This technique is used to observe the projected potential of InP along the (001) direction, to measure the thickness of InP crystals, and to identify In and P atom columns from a high-resolution electron hologram

Inhibition of porcine pancreatic alpha-amylase (1,4-alpha-D-glucan glucanohydrase) [EC 3.2.1.1] with maltotriitol (G3OH) and 4-phenylimidazole was investigated by using maltohexaitol (G6OH) and p-nitrophenyl-alpha-D-maltoside (G2PNP) as substrates. When G6OH was the substrate, both G3OH and 4-phenylimidazole behaved as competitive inhibitors. On the other hand, when G2PNP was the substrate, G3OH behaved as a competitive inhibitor, whereas 4-phenylimidazole behaved as a non-competitive inhibitor. Further inhibition study in the presence of both G3OH and 4-phenylimidazole, with G6OH as the substrate, showed that the two inhibitors compete with each other for the active site of the enzyme. Based on a consideration of the productive (reactive) binding modes of G2PNP and G6OH, and a nonproductive (nonreactive) binding mode of G2PNP, it is suggested that the binding sites of the two inhibitors may be partially overlapping around the catalytic site of the enzyme and that the rest of the binding site of each inhibitor lies along the substrate binding cleft of the enzyme.

(1992). Isolation and Structural Elucidation of the Major Genuine Soybean Saponin. Bioscience, Biotechnology, and Biochemistry: Vol. 56, No. 1, pp. 142-143.

1. The activities of 2-oxoglutarate dehydrogenase, transketolase, thiamine pyrophosphokinase and thiamine triphosphatase and the concentrations of thiamine phosphates were almost the same between rat extensor digitorum longus and soleus muscles at 2 weeks of age. 2. These enzyme activities changed after 3 weeks of age in a different way depending on the muscle phenotype. 3. Thiamine diphosphate level and the activity of 2-oxoglutarate dehydrogenase increased only in soleus muscle and thiamine triphosphate level increased only in extensor digitorum longus during development.

1. Thiamine triphosphatase activity in particulate fraction, but not in soluble, of rat skeletal muscle was stimulated by several anions. 2. The stimulative effect of anions was dependent on pH of reaction medium and was reversible. 3. The activities of ATPase in rat muscle particulate preparation and thiamine triphosphatase in the brain were inhibited by the anions.

We studied the capping growth process of an InGaAs on high-density InAs/GaAs(001) quantum dots (QDs) with a bimodal size distribution grown by molecular beam epitaxy. Atomic force microscopy and reflection high energy electron beam diffraction observations revealed that the bimodal size distribution is composed of larger QDs with {110} facets and smaller QDs with {136} facets. In addition, it was found that, during the InGaAs capping growth, the height of larger QDs decreased, while the height of smaller QDs increased due to the incorporation of indium adatoms. As a result, the size fluctuation of the QDs was suppressed as compared to GaAs capping growth. We achieved a narrow photoluminescence (PL) linewidth of 24 meV and an enhanced PL peak intensity.

Zymomonas mobilis IFO 13 756 could take up galactose and metabolize it so that galactonic acid accumulated in the culture fluid. The oxidation of galactose was catalysed by a membrane-associated enzyme, which could be solubilized with a mixture of 1% Triton X-100 and 1 M: KC1. The strain also accumulated galactitol as a minor metabolite. To confer the ability to utilize galactose on Z. mobilis, a Gal(+) recombinant plasmid, pZG13, was constructed by the insertion of the galETK genes of Escherichia coli immediately downstream of the Z. mobilis promoter gene in pZA22, the plasmid introduced into a Zymomonas strain. Uridine diphospho-glucose 4-epimerase coded in the galE gene was expressed in Z. mobilis carrying pTG13. The recombinant strain could produce a small amount of ethanol from galactose.

We have visualized DNA molecules in solution by Fraunhofer in-line electron holography. So far, applications of the technique at high resolution have been limited to gold particles. This report is the first application of Fraunhofer in-line electron holography to a nanometer-sized biological specimen.The phase shift of an electron beam due to DNA is so small, estimated as only 2π/33 for 100-keV electrons, that it has been only possible to visualize them with either heavy-atom staining or metal shadowing techniques. However, these methods give only low-resolution images of the molecules, and worst of all, preservation of specimens is poor. The frozen-hydrated specimen preparation improved the preservation of specimen, and images of naturally supercoiled DNA in solution have been visualized using underfocusing of 1-2 μm. Such a defocusing is large enough for such a small object as DNA (diameter ∽ 2 nm) to be in a Fresnel condition;z≈ a2/λ, wherezis the defocusing, a is the diameter of an object, andλis the wavelength.

Treatment of [RuCl2(COD)]n with (R)-p-MeO-BINAP (p-MeO-BINAP = 2,2‘-bis(bis(p-methoxyphenyl)phosphino)-1,1‘-binaphthyl) in toluene in the presence of triethylamine afforded the anionic dinuclear complex [NH2Et2][{RuCl((R)-p-MeO-BINAP)}2(μ-Cl)3] ((R)-2), whose structure has been determined by an X-ray crystallographic study. Complex 2 is an efficient catalyst for asymmetric hydrogenation of functionalized olefins and ketones.

Electron interference micrographs of a single magnetic-domain particle of barium ferrite are obtained experimentally and theoretically. In the experiment, a hologram of the particle with size about 1 µ m is recorded on film and the phase distribution of the object electron wave is reconstructed digitally by the Fourier transform method. In the theoretical calculation, the phase distribution is obtained by integrating the vector potential around the particle along the electron path, assuming that the particle is spherical and is in the single-domain state. The above two phase distributions are converted to interference micrographs where magnetic flux lines are observed, and compared with each other. The theoretically calculated result agrees well with the experimental one, proving that the particle is in the single-domain state.

Myosins purified from cardiac (porcine heart) and smooth (chicken gizzard) muscles were modified with 2,4,6-trinitrobenzenesulfonate (TNBS) and the effects on the kinetic properties of myosin ATPase [EC 3.6.1.3] were studied. The following results were obtained. 1. About 0.5 mol of TNBS per mol of myosin head was incorporated rapidly, irrespective of the presence of PP1 (2mM), into both types of myosin studied. 2. The size of the initial burst of P1 liberation for both myosins was found to be 0.5--0.6 mol/mol head. 3. The rapid incorporation of TNBS into cardiac muscle myosin was accompanied by a rapid decrease in the size of the initial P1 burst, and it was completely lost after modification for 20 min. However, smooth muscle myosin retained its P1 burst. 4. The EDTA (K+)-ATPase activity of both myosins modified in the presence or absence of PP1 decreased sharply with incorporation of TNBS. 5. Superprecipitation and ATPase activity of reconstituted actomyosin from cardiac myosin and skeletal F-actin decreased only after 10 min of modification with TNBS in the absence of PP1. 6. The spectra of TNP bound to myosins from cardiac and smooth muscles were unchanged by the addition of PP1. The above findings are compared with those previously obtained for skeletal muscle myosin [Miyanishi, T., Inoue, A., & Tonomura, Y. (1979) J. Biochem. 85, 747--753], and the structural and functional differences among the myosins derived from skeletal, cardiac, and smooth muscles are discussed.

Na+, K+-dependent ATPase [EC 3.6.1.3] was purified from porcine kidney by the method of Lane et al. [(1973) J. Biol. Chem. 248, 7197-7200] with slight modifications [Yamaguchi, M. & Tonomura, Y., (1979) J. Biochem. 86, 509-523]. The amounts of a phosphorylated intermediate (EP) and ouabain bound to the enzyme during the ATPase reaction were measured in 2.1 mM MgCl2 and various concentrations of NaCl and KCl at pH 7.5 and 20 degrees C. In presence of NaCl and the absence of KCl, the molar ratio of the amounts of EP and bound ouabain was 1 : 2. In the presence of both NaCl and KCl, it was 1 : 1. In both cases, the amount of bound ouabain was equal to that of EP in the absence of ouabain. These findings suggest that the functional unit of the transport ATPase is a dimer.

The amount of Ca2+ bound to the Ca2+,Mg2+-dependent ATPase of deoxycholic acid-treated sarcoplasmic reticulum was measured during ATP hydrolysis by the double-membrane filtration method [Yamaguchi, M. & Tonomura, Y. (1979), J. Biochem. 86, 509--523]. The maximal amount of phosphorylated intermediate (EP) was adopted as the amount of active site of the ATPase. In the absence of ATP, 2 mol of Ca2+ bound cooperatively to 1 mol of active site with high affinity and were removed rapidly by addition of EGTA. AMPPNP did not affect the Ca2+ binding to the ATPase in the presence of MgCl2. Under the conditions where most EP and ADP sensitive at steady state (58 microM Ca2+, 50 microM EGTA, and 20 mM MgCl2 at pH 7.0 and 0 degrees C), bound Ca2+ increased by 0.6--0.7 mol per mol active site upon addition of ATP. The time course of decrease in the amount of bound 45Ca2+ on addition of unlabeled Ca2+ + EGTA was biphasic, and 70% of bound 45Ca2+ was slowly displaced with a rate constant similar to that of EP decomposition. Similar results were obtained for the enzyme treated with N-ethylmaleimide, which inhibits the step of conversion of ADP-sensitive EP to the ADP-insensitive one. Under the conditions where most EP was ADP insensitive at steady state (58 microM Ca2+, 30 microM EGTA, and 20 mM MgCl2 at pH 8.8 and 0 degrees C), the amount of bound Ca2+ increased slightly, then decreased slowly by 1 mol per mol of EP formed after addition of ATP. Under the conditions where about a half of EP was ADP sensitive (58 microM Ca2+, 25 microM EGTA, and 1 mM MgCl2 at pH 8.8 and 0 degrees C), the amount of bound Ca2+ did not change upon addition of ATP. These findings suggest that the Ca2+ bound to the enzyme becomes unremovable by EGTA upon formation of ADP-sensitive EP and is released upon its conversion to ADP-insensitive EP.

Electron holograms of H-Nb2O5 are formed from about 150 diffracted waves used to reconstruct high-resolution images. A niobium atom arrangement in H-Nb2O5 is observed in an aberration-corrected phase image. A hologram with 9.3-pm carrier fringes is also formed. It is the finest carrier fringe spacing hologram ever reported.

A 350 kV electron microscope equipped with a magnetic-field-superimposed field emission gun has been developed. A 0.055 nm-spacing lattice image in a gold thin film has been successfully observed with this electron microscope under the condition of 300 kV electron accelerating voltage.

Solubilized sarcoplasmic reticulum (SSR) was prepared by solubilizing fragmented sarcoplasmic reticulum (FSR) with a nonionic detergent (C12E8) then displacing the detergent with Tween 80, using a DEAE-cellulose column. The UV absorption of SSR decreased reversibly at about 286 and 292 nm on removal of free Ca2+ ions, while no change in the fluorescence spectrum was detectable. On the other hand, the fluorescence intensity of FSR decreased 3-4% on removal of free Ca2+ ions, as previously reported by Dupont [(1976) Biochem. Biophys. Res. Commun. 71, 544-550]. The UV absorption of FSR increased reversibly at about 270-280 nm on removal of free Ca2+ ions, but the rate of the change was very slow (k = about 0.1 min-1).

Fragmented sarcoplasmic reticulum (FSR) from rabbit skeletal muscle was passively loaded with 45Ca2+. Its Ca2+-induced Ca2+ release was measured in the presence of 0.1 M KCl and 5 mM MgCl2 at 0 degrees C by Millipore filtration. The following results were obtained. 1. The amounts of Ca2+-induced Ca2+ release from heavy SR, light SR, and unfractionated SR were 80, 20, and 60% of the amounts of preloaded Ca2+, respectively. Therefore, the experiments were carried out with unfractionated FSR. 2. The Ca2+-induced Ca2+ release from FSR was inhibited by procaine, but unaffected by caffeine and trifluoperazine. The rate of Ca2+ release decreased markedly with decreasing pH. 3. Various adenine nucleotides (ATP, AMPPNP, ADP, AMP) accelerated the Ca2+ release, and the accelerating effect was reversible. CTP had no effect on the release, but inhibited the accelerating effect of AMPPNP. 4. In the presence of 15 microM external free Ca2+, the final amount of the Ca2+ release was unaffected. The rate of Ca2+ release was markedly increased by AMP; the dependence of the rate on AMP concentration followed a Michaelis-Menten type equation with a Hill coefficient of 1 and an apparent affinity for AMP of about 2 mM. 5. In the presence of AMP, the amount of Ca2+ released increased, while the relative rate was unaffected by increasing the external Ca2+ concentration. The final amount released increased from 0 to 60% of the amount of preloaded Ca2+ by increasing the free Ca2+ concentration from 0.06 to 0.24 microM. The effect of external Ca2+ on the release was reversible. 6. The ratio between the amount of preloaded Ca2+ and that of Ca2+ release was independent of the Ca2+ concentration used for preloading. Furthermore, the dependence of the final amount of Ca2+-induced Ca2+ release on external Ca2+ was unaffected by internal Ca2+.