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Biomedical subjects

H Akutsu

Publications and source records attributed to H Akutsu.

At least 127 records · Page 7Linked to original sources
PubMed1984

Photochemically induced dynamic nuclear polarization study on microsomal NADPH-cytochrome P-450 reductase.

Sedimentation equilibrium experiments with NADPH-cytochrome P-450 reductase showed that increasing 1-O-n-octyl-beta-D-glucopyranoside levels promoted disaggregation of the flavoprotein. The reductase was monomeric at a molar ratio of detergent to protein above 10(3). Addition of N3-carboxymethyllumiflavin to the flavoprotein in the presence of 1-O-n-octyl-beta-D-glucopyranoside results in photochemically induced dynamic nuclear polarization (CIDNP) signals in the aromatic region. The CIDNP spectrum of the holoprotein shows sharp resonances due to histidine residues. On removal of FMN from the protein, CIDNP signals originating from a tyrosine residue appeared, suggesting that the tyrosine residue is exposed to solvent after the depletion of FMN. However, this tyrosine residue appears to become inaccessible to the external dye after full incubation of FMN-depleted reductase with FMN. This suggests that the tyrosine residue could be located in the vicinity of the FMN-binding domain which constitutes the active center of the reductase.

Animals↗
PubMed1984

High resolution NMR study of CAP binding site 22mer in H2O solution.

High resolution proton NMR were measured for the deoxyoligonucleotide 22mer duplex corresponding to the CAP (catabolite gene activator protein) binding site of lac promotor. The spectra in the lower field region than the water resonance were taken with the time-shared Redfield pulse method by using a JEOL 500 MHz NMR spectrometer. In the imino proton region 18 peaks were separately observed, but the area intensity at 10 degrees C corresponds to 20 protons. By selective irradiation at each peak position NOEs (nuclear Overhauser effects) were observed between the imino and adenine C2H protons and between imino proton themselves. By tracing sequential NOE train carefully, 17 imino proton signals could be unambiguously assigned to each base pair except five AT base pairs at terminals. With the elevation of temperature the peaks showed gradual broadening and disappeared, which indicates the stepwise base pair opening of the duplex. Referring to the above peak assignments it can be concluded that GC20 and AT4 pairs close to terminals relax first and the base pair opening proceeds toward central GC13 and 14.

Base Sequence↗
PubMed1983

Behavior of the imino protons of the lambda OR3 17mer studied by high resolution NMR.

Imino proton resonances of lambda OR3 17mer were observed with time-shared Redfield pulse method by using a JEOL 500 MHz NMR instrument. They show gradual broadening and disappearance with the elevation of temperature indicating the stepwise melting of the duplex. By the selective irradiation at each peak position nuclear Overhauser effects were observed between the imino and adenine C2H protons and between imino protons themselves. Combining these data, fifteen imino proton resonances could be assigned to each base pair except two terminal AT base pairs. Based on the assignment it can be said that AT rich regions near the terminal melt first, followed by the melting of the inside GC rich core. The two AT base pairs in the middle of the GC core are resistant to heat. Spin lattice relaxation times were also observed and the results are consistent with the melting profile.

Bacteriophage lambda↗
PubMed1983

Two-dimensional echo-cardiographic estimation of left atrial volume and volume load in patients with congenital heart disease.

To estimate the left atrial volume (LAV) and pulmonary blood flow in patients with congenital heart disease (CHD), we employed two-dimensional echocardiography (TDE). The LAV was measured in dimensions other than those obtained in conventional M-mode echocardiography (M-mode echo). Mathematical and geometrical models for LAV calculation using the standard long-axis, short-axis and apical four-chamber planes were devised and found to be reliable in a preliminary study using porcine heart preparations, although length (10%), area (20%) and volume (38%) were significantly and consistently underestimated with echocardiography. Those models were then applied and correlated with angiocardiograms (ACG) in 25 consecutive patients with suspected CHD. In terms of the estimation of the absolute LAV, accuracy seemed commensurate with the number of the dimensions measured. The correlation between data obtained by TDE and ACG varied with changing hemodynamics such as cardiac cycle, absolute LAV and presence or absence of volume load. The left atrium was found to become spherical and progressively underestimated with TDE at ventricular endsystole, in larger LAV and with increased volume load. Since this tendency became less pronounced in measuring additional dimensions, reliable estimation of the absolute LAV and volume load was possible when 2 or 3 dimensions were measured. Among those calculation models depending on 2 or 3 dimensional measurements, there was only a small difference in terms of accuracy and predictability, although algorithm used varied from one model to another. This suggests that accurate cross-sectional area measurement is critically important for volume estimation rather than any particular algorithm involved. Cross-sectional area measurement by TDE integrated into a three dimensional equivalent allowed a reliable estimate of the LAV or volume load in a variety of hemodynamic situations where M-mode echo was not reliable.

Adolescent↗
PubMed1982

Local anesthetics and divalent cations have the same effect on the headgroups of phosphatidylcholine and phosphatidylethanolamine.

The effects of the local anesthetic dibucaine on the membrane headgroup conformations of phosphatidylcholine and phosphatidylethanolamine were determined using 2H- and 31P-NMR. The size of the deuterium quadrupole splittings of the two methylene segments of the choline and ethanolamine groups changed dramatically and the 31-phosphorus chemical shift anisotropy of the phosphatidylcholine headgroup decreased by about 7 ppm in the presence of local anesthetic. The quadrupole splittings of the 3-glycerol and choline methyl segments were relatively insensitive to the addition of dibucaine. The headgroup data for dibucaine addition paralleled similar data for the addition of various cations. These NMR results agree with the previous observation that these drugs displace calcium from phospholipids. The effects of this local anesthetic on these headgroups were distinctly different from the changes induced by cholesterol, heat and the general anesthetic chloroform.

Cations, Divalent↗
PubMed1982

Structure of the lambda tof repressor protein in solution. Heat stability and its relation to binding ability to DNA.

The lambda tof repressor protein was purified from E. coli cells retaining lambda dv plasmids by applying DNA-cellulose chromatography. 3H-labeled lambda dv and lambda imm21dv DNA, carrying and lacking lambda operators, respectively, were prepared and the binding activity of the lambda tof protein to the DNA was examined. Non-specific binding to lambda imm21dv DNA is completely lost at 30 degrees C, whereas specific binding to the DNA carrying the operators is retained even above 40 degrees C. The conformation of the lambda tof protein was analysed by means of circular dichroism and 1H-NMR spectra. The change in the molar ellipticity at 222 nm vs. temperature in CD spectra indicated a transition between two states with Tm at 42 degrees C. The 360 MHz 1H-NMR spectra revealed the presence at 20 degrees C of another change in local conformation of interaction which was not detected by the CD spectra. 1H-NMR also indicated the coexistence of thermal transitions with exchange rates faster and slower than the NMR time scale at about 50 degrees C, which is explained by the presence of domain structures. The NMR titration curve of the His residue gave a normal pK value showing its location on the surface of the protein. These conformational behaviors are well correlated to the specific and non-specific DNA binding activity of the lambda tof protein. The assignments of 1H resonance signals to some specific residues, including His 35 and Tyr 26, were established. It will be useful to determine the tof-DNA interaction.

Bacterial Proteins↗
PubMed1981

Direct determination by Raman scattering of the conformation of the choline group in phospholipid bilayers.

For clarification of the assignment of the vibrational modes of the choline group, Raman spectra of choline iodides selectively deuterated at three different positions were investigated. The isotope shifts of the C-N stretching vibrations suggested that they are conformation sensitive. When the Raman spectra of choline chloride, carbamoylcholine iodide, carbamoylcholine chloride, and methoxycarbonylcholine iodide are compared with the crystal structures of these compounds, a correlation between the vibrational frequency and the conformation of the O-C-C-N+ backbone could be established. The Raman bands attributed to the "totally" symmetric stretching (v1) and symmetric stretching vibrations (v2) of the C-N bonds of the quaternary ammonium group appeared at about 720 cm-1 and about 870 cm-1, respectively, for the gauche conformation of the O-C-C-N+ backbone, and in the trans conformation, they shifted to about 770 cm-1 (v1) and about 910 cm-1 (v2), respectively. On the basis of this correlation and from measurements of phosphatidylcholine and sphingomyelin bilayers, it was concluded that most of the choline groups in both bilayers take the gauche conformation not only in the solid state but also in the gel and liquid-crystalline states. These data represent the first direct evidence that a gauche conformation for the O-C-C-N+ bond is preferred in the gel and liquid-crystalline states. These key bands, especially the v1 band, are a powerful tool to study the conformation of the choline group in situ not only in the membrane field but also in the neuroscience in connection with acetylcholine.

Choline↗
PubMed1980

Evidence for phase separation in the membrane of an osmotically stabilized fatty acid auxotroph of E. coli and its biological significance.

1. An unsaturated fatty acid auxotroph of Escherichia coli accumulated a high content of saturated fatty acids in its membrane when it was cultured under osmotically stabilized conditions. The physicochemical properties of the phospholipid extracts and of the membrane fraction from the cells were investigated by means of proton magnetic resonance, infrared spectroscopy and differential scanning calorimetry. 2. Physicochemical studies indicate that the phospholipid bilayers in the membranes exhibit at least two phase transitions, a minor one at approx. 19 degrees C and a major one at approx. 43 degrees C. Between the two temperatures, gel and liquid crystalline domains co-exist. Moreover, even in the gel state, phospholipids seem to segregated into domains containing different proportions of unsaturated fatty acids. 3. The Arrhenius plot of beta-galactoside transport rates is biphasic. The inflection point is at 22 degrees C. This means that the appearance of the fluid region in the bilayer at approx. 19 degrees C is important in the activation of membrane transport.

Biological Transport↗
PubMed1980

Structure and synthesis of a lipid-containing bacteriophage. Studies on the structure of the bacteriophage PM2 nucleocapsid.

The nucleocapsid of bacteriophage PM2, prepared according to Schäfer et al. [Eur. J. Biochem. 92, 579-588 (1978)], was studied by biochemical and biophysical methods. It was not possible to isolate the lipid-free nucleocapsid. More than 95% of the lipids were associated with the nucleocapsid. The asymmetric distribution of phospholipids across the viral membrane was retained in the nucleocapsid since less than 10% of the phosphatidylethanolamine was accessible to the non-penetrable membrane probe, 2,4,6-trinitrobenzenesulfonate. Micro-dissection of the nucleocapsid with thermolysin demonstrated the asymmetric orientation of core proteins across the nucleocapsid membrane. Protein III was embedded deeply in the lipid bilayer and about 20% of the molecule extended to the exterior. Protein IV interacts with PM2 DNA and is partially in the inner leaflet of the bilayer. Small-angle X-ray scattering studies on the nucleocapsid enabled us to localize the lipid bilayer structure at radii from 20.5 nm to 24.0 nm.

Bacteriophages↗
PubMed1979

Capsimycin, a new antibiotic. I. Production, isolation and properties.

Capsimycin is a new antifungal antibiotic produced by a strain of Streptomyces sp. C 49--87. The active substance in the fermented broth was isolated by solvent extraction followed by silica gel column chromatography. The antibiotic melts at 186 degrees C (decomp.) and has a molecular formula C30H40N2O6. It exhibits marked inhibitory activity against Phytophthora capsici (Leaf blight disease of cucumber) and Pythium debaryanum (Damping-off disease of cucumber).

Animals↗