PubMed Health⌕ Search

Biomedical subjects

K Nagayama

Publications and source records attributed to K Nagayama.

At least 19 recordsLinked to original sources
PubMed1992

1H NMR studies of deuterated ribonuclease HI selectively labeled with protonated amino acids.

Two-dimensional (2D) 1H NMR experiments using deuterium labeling have been carried out to investigate the solution of ribonuclease HI (RNase HI) from Escherichia coli (E. coli), which consists of 155 amino acids. To simplify the 1H NMR spectra, two fully deuterated enzymes bearing several protonated amino acids were prepared from an RNase HI overproducing strain of E. coli grown in an almost fully deuterated medium. One enzyme was selectively labeled by protonated His, Ile, Val, and Leu. The other was labeled by only protonated His and Ile. The 2D 1H NMR spectra of these deuterated RNase HI proteins, selectively labeled with protonated amino acids, were much more simple than those of the normally protonated enzyme. The simplified spectra allowed unambiguous assignments of the resonance peaks and connectivities in COSY and NOESY for the side-chain protons. The spin-lattice relaxation times of the side-chain protons of the buried His residue of the deuterated enzyme became remarkably longer than that of the protonated enzyme. In contrast, the relaxation times of the side-chain protons of exposed His residues remained essentially unchanged.

Amino Acid Sequence↗
PubMed1992

Noninvasive assessment of cardiomyopathy development with simultaneous measurement of topical 1H- and 31P-magnetic resonance spectroscopy.

Background. It might be possible to estimate the metabolic derangement of cardiac muscle by topical nuclear magnetic resonance spectroscopy (MRS) in vivo without killing the animal. Methods and Results. By use of topical 1H- and 31P-MRS focused on the heart of Syrian hamsters with or without cardiomyopathy (CM; BIO 14.6 strain), the chemical constituents were measured in vivo nondestructively and repetitively at several stages of development of CM. A phantom experiment and two-dimensional plot of chain methylenes (CH2) of lipid/water ratio by 1H-MRS versus creatine phosphate (CP)/[beta-P]ATP ratio by 31P-MRS indicated that signal cross talk from the adjacent organs was negligible. Even before the onset of clinical or pathological manifestation of CM (7 weeks after birth), CH2/water ratio by 1H-MRS was lower in the CM group (7.3 +/- 0.7%) than in control (11.8 +/- 2.0%, p less than 0.05), and it decreased further at the hypertrophic stage (17 weeks, 4.1 +/- 0.7%, p less than 0.05) and the congestive stage (27 weeks, 4.3 +/- 0.9%, p less than 0.05). In contrast, the CP/[beta-P]ATP ratio by 31P-MRS started to decrease at the hypertrophic stage (1.90 +/- 0.18 versus 2.52 +/- 0.24, p less than 0.05) and decreased further at the congestive stage to 1.53 +/- 0.18 (p less than 0.01). These in vivo MRS data were confirmed by both biochemical assay and in vitro MRS analysis in heavy water after the animals were killed. Conclusions. A combination of topical 1H-MRS and 31P-MRS in vivo is promising for the noninvasive and sensitive assessment of cardiac muscle metabolism. Comparison of these MRS studies and biochemical analysis suggested not only the modification of water, lipid, CP, or ATP contents but also the reduction of flexibility or fluidity of lipids in cardiomyopathic heart.

Adenosine Triphosphate↗
PubMed1991

Assignments of backbone 1H, 13C, and 15N resonances and secondary structure of ribonuclease H from Escherichia coli by heteronuclear three-dimensional NMR spectroscopy.

The assignments of individual magnetic resonances of backbone nuclei of a larger protein, ribonuclease H from Escherichia coli, which consists of 155 amino acid residues and has a molecular mass of 17.6 kDa are presented. To remove the problem of degenerate chemical shifts, which is inevitable in proteins of this size, three-dimensional NMR was applied. The strategy for the sequential assignment was, first, resonance peaks of amides were classified into 15 amino acid types by 1H-15N HMQC experiments with samples in which specific amino acids were labeled with 15N. Second, the amide 1H-15N peaks were connected along the amino acid sequence by tracing intraresidue and sequential NOE cross peaks. In order to obtain unambiguous NOE connectivities, four types of heteronuclear 3D NMR techniques, 1H-15N-1H 3D NOESY-HMQC, 1H-15N-1H 3D TOCSY-HMQC, 13C-1H-1H 3D HMQC-NOESY, and 13C-1H-1H 3D HMQC-TOCSY, were applied to proteins uniformly labeled either with 15N or with 13C. This method gave a systematic way to assign backbone nuclei (N, NH, C alpha H, and C alpha) of larger proteins. Results of the sequential assignments and identification of secondary structure elements that were revealed by NOE cross peaks among backbone protons are reported.

Amino Acid Sequence↗
PubMed1991

Membrane conductance of an electroporated cell analyzed by submicrosecond imaging of transmembrane potential.

Transmembrane potential was induced in a sea urchin egg by applying a microsecond electric pulse across the cell. The potential was imaged at a submicrosecond time resolution by staining the cell membrane with the voltage-sensitive fluorescent dye RH292. Under moderate electric fields, the spatial distribution of the induced potential as well as its time dependence were in accord with the theoretical prediction in which the cell membrane was regarded as an insulator. At higher field intensities, however, the potential apparently did not fully develop and tended to saturate above a certain level. The saturation is ascribed to the introduction of a large electrical conductance, in the form of aqueous openings, in the membrane by the action of the induced potential (electroporation). Comparison of the experimental and theoretical potential profiles indicates that the two regions of the membrane that opposed the electrodes acquired a high membrane conductance of the order of 1 S/cm2 within 2 microseconds from the onset of the external field. The conductance was similar in the two regions, although permeability in the two regions of the membrane long after the pulse treatment appeared quite different.

Animals↗
PubMed1991

Image analysis by electron microscopy of two-dimensional crystals developed on a mercury surface of chaperonin from Thermus thermophilus.

Two-dimensional crystals of functional chaperonin molecules, which are protein complexes of cpn60 and cpn10, isolated from Thermus thermophilus were prepared on a mercury surface under oxygen atmosphere and were observed by electron microscope after transferring them to carbon coated specimen grids. The crystals showed the hexagonal lattice with unit cell dimensions of a = b = 12.4 nm and gamma = 120 degrees. The averaged image at a 3 nm resolution of the chaperonin has a doughnut-like shape which has seven peripheral masses and a central cavity. Preincubation of the chaperonin with MgATP changed the mobility in non-denaturating PAGE but did not cause distinguishable change of shape. The location of cpn10 in the chaperonin molecule is discussed.

Bacterial Proteins↗
PubMed1991

[A case of lupus nephritis with hyporeninemic hypoaldosteronism].

We report a case of 67-year-old woman with systemic lupus erythematosus presenting hyporeninemic hypoaldosteronism. She admitted because of anasarca in March, 1990. She manifested nephrotic syndrome, and hyperkalemia and hyperchloremic metabolic acidosis. The hyperkalemia was disproportionate to the degree of renal insufficiency. Basal levels of plasma renin activity and plasma aldosterone concentration were low. Renal tubular function studies revealed normal hydrogen ion secretion. Renal biopsy demonstrated diffuse proliferative lupus nephritis and prominent interstitial cell infiltration. There was no vasculitis of glomerular vascular poles. After treatment of lupus nephritis with prednisolone, levels of plasma renin activity and plasma aldosterone concentration were elevated. Hyperkalemia and metabolic acidosis were normalized and renal function improved. We conclude that the heperkalemia and metabolic acidosis could be attributed to hyporeninemic hypoaldosteronism.

Acidosis↗
PubMed1990

NMR characteristics of intracellular K in the rat salivary gland: a 39K NMR study using double-quantum filtering.

Intracellular K of the perfused rat mandibular salivary gland was measured by 39K NMR spectroscopy at 8.45 T. Multiple-quantum NMR arising from multiple-exponential decay was used to eliminate the resonance due to extracellular K in the perfused gland at 25 degrees C. The resonance due to intracellular K consisted of two Lorentzian signals stemming from the [spin 1/2 to -1/2] coherence (sharp resonance) and the [spin -1/2 to -3/2], [spin 3/2 to 1/2] coherences (broad resonance). The transverse relaxation time (T2) corresponding to the [spin 1/2 to -1/2] coherence was ca. 2.5 ms, and that corresponding to the [spin -1/2 to -3/2], [spin 3/2 to 1/2] coherences was ca. 0.4 ms. The relaxation time of the double-quantum coherence of rank 3 (originating from product operators like Ix2Iz) was determined to be ca. 0.2 ms. These results suggest the possibility of the presence of a single homogeneous population of intracellular K with a correlation time of ca. 2.5 x 10(-8) s and a quadrupolar coupling constant of ca. 1.4 MHz.

Animals↗
PubMed1990

Combination of heteronuclear 1H-15N and 1H-13C three-dimensional nuclear magnetic resonance experiments for amide-directed sequential assignment in larger proteins.

A new strategy for the sequential assignment of backbone proton resonances in larger proteins involving a unique combination of four types of heteronuclear three-dimensional (3D) NMR spectroscopies is reported. This method relies on the uniform labeling of amide nitrogens with 15N and of alpha-carbons with 13C. Heteronuclear 1H-15N TOCSY-HMQC and NOESY-HMQC experiments can reveal connections between cross-peaks arising from the NHi-C alpha Hi-1 and NHi-C alpha Hi connectivities in the finger-print region in in general. They also specifically reveal the sequential amide-amide connectivities among the amide cross-peaks for the alpha-helices. Heteronuclear 1H-13C HMQC-TOCSY and HMQC-NOESY experiments can reveal connections between cross-peaks arising from the NHi-C alpha Hi and NHi+1-C alpha Hi connectivities in the finger-print region in general. The combination of the two sets of results reveals the complete unambiguous sequential connection of cross-peaks for the proton resonances in the peptide backbone. The application of the new strategy is reported for a protein, ribonuclease H, with a molecular weight of 17.6 kDa.

Amino Acids↗
PubMed1990

Quantitative chemical analysis of nickel-chromium dental casting alloys.

Twenty-nine brands of dental casting nickel-chromium alloys made in Japan for small castings were analyzed by electron probe X-ray microanalyzer. Nickel-chromium alloys for metal-ceramic application were composed primarily of nickel, chromium, and molybdenum with the exception of one brand. Of the nickel-chromium alloys for inlay, crown, and bridgework applications, 11 of the 22 alloys were up to the standard of the Ministry of Welfare specifications. And additive metal elements of these alloys were molybdenum, iron, copper, manganese, aluminum, silicon, tin, indium, silver, titanium, and gallium.

Chromium Alloys↗
PubMed1990

[Influence of metal elements in Pd alloys on the bonding to porcelain].

The influence of metal elements in three Pd alloys on the bonding to porcelain was examined. The high-temperature oxidation behavior of metal elements in alloys, structure and morphology of oxidation zones on the alloy surface and the appearance of bonding between alloy and fused porcelain at the interface were studied by ESCA and EPMA. From ESCA analysis, metal oxides such as PdO, SnO2, In2O3, CuO or Cu2O, Sb2O3 or Sb2O5 were formed on the surface of the alloy after degassing (preheating) and heating following casting. The amount of metal elements concentrated on the alloy surface was measured by their characteristic X-rays using area analysis of EPMA under the technical conditions of A) Acc. voltage 10 kV, Abs. current 40 nA, and B) Acc. voltage 20 kV, Abs. current 10 nA. Consequently, the amount of additive metal elements concentrated on the alloy surface which was detected in condition A) was larger than that detected in condition B). However, the amount of main element (Pd) concentrated on the alloy surface which was detected in condition A) was smaller than that detected in condition B). From the EPMA observation of the interface between alloy and fused porcelain, cross-section of alloys showed that the additive metal elements were oxidized in the alloy matrix, forming an internal oxide layer or particles. The structure and morphology of oxidation zones showed a significant difference between degassed and not-degassed alloys. Cross-sections inclined approximately 5 degrees 43' to the fusing boundary were also analyzed to enlarge the oxidation zones. This analytical technique effectively revealed the structure and morphology of oxidation zones. In conclusion, not only additive metal elements but also the main element at the surface of alloys play important roles on the bonding to porcelain. Degassing of the alloy was considered to an essential procedure to obtain the adequate bonding to porcelain.

Dental Bonding↗
PubMed1990

[Crystalline glass ceramics model teeth].

The feeling of hardness, stickiness and the rate of cutting during the preparation of a tooth were compared using an extracted tooth, melamine resin and crystalline glass ceramics model tooth (Bioram M) which are used in dental education and practice. Diamond burs were used with an air-turbine handpiece in this experiment. The results indicated that Bioram M had the desirable characteristics of a model tooth and there was a strong resemblance feeling of cutting between Bioram M and a natural vital tooth.

Ceramics↗
PubMed1990

[Studies on titanium casting. (1) Influence of the mold temperature on titanium castings].

Pure titanium and titanium alloys have excellent biocompatibility, excellent corrosion tarnish resistance, a quarter of the specific gravity of gold base alloys and suitable mechanical properties for dental clinical use. However, titanium has some undesirable characteristics. For example, it has a high melting temperature and chemical reactivity at high temperatures. The investigate influence of the mold temperature on castability and mechanical properties of the titanium castings was study. The castability was found to be good at various mold temperatures, but the higher the mold temperature during casting, the greater the adhesive phenomenon between titanium surface and the investment. From the EPMA observation of the surface layer on the titanium castings, an Si- rich layer whose thickness increased considerably with increasing temperature was observed. The Vickers hardness in the surface layer was greater than that in the inner part. At a higher mold temperature during casting the thickness of the high Vickers hardness layer was increased. There was a relationship between the Vickers hardness value and the thickness of Si rich layer. In the tensile strength test of titanium castings, elongation values decrease and tensile strength values increased with the rise in mold temperature.

Dental Alloys↗
PubMed1989

Hexagonal structure of two-dimensional crystals of the alpha 3 beta 3 complex thermophilic ATP synthase.

The highly dissociable alpha 3 beta 3 subunit complex (Mr = 319,582) of thermophilic ATP synthase was crystallized on a mercury surface under oxygen. The two-dimensional crystal was compared with that of TF1 (Mr = 385,351, alpha 3 beta 3 gamma delta epsilon subunit complex) by means of computer image processing. The crystals showed the same hexagonal lattice (a = b = 10 nm), despite the difference in their molecular weights. The color images of the two protein molecules were also hexagonal. However, there was an open hole in the image of the alpha 3 beta 3 complex, where small subunits (gamma, delta, and epsilon) of TF1 may have been located. The structure of this heterohexamer is consistent with that deduced from other physical parameters.

Crystallization↗
PubMed1989

The occurrence of intramitochondrial Ca2+ granules in valproate-induced liver injury.

Two epileptic patients treated with anticonvulsants of valproic acid, phenobarbital, nitrazepam, and diphenylhydantoin developed anorexia, convulsions, and unconsciousness. The liver biopsy samples showed degenerated hepatocytes containing enlarged mitochondria with a distorted matrix. The matrix granules in the mitochondria were enlarged in size as compared with those of chronic hepatitis (Po = 0.0009). X-ray microanalysis has revealed the presence of calcium in large mitochondrial matrix granules. Both patients were thought to have valproic acid intoxication because they recovered quickly after the withdrawal of valproic acid.

Calcium↗
PubMed1989

[Application of Holter electrocardiography to exercise testing for the group examination of school children: comparison between obese and normal children].

Application of Holter electrocardiography to an exercise test for heart function is useful in the group examination of school children. The circulatory responses to a step test among obese boys and girls and control were as follows: 1. Four cases of arrhythmia were detected, of whom 2 cases were discovered during exercise. ST-segment depression of 0.1 mV or more during and after exercise was recognized in 29 cases, 27 of the junctional- and 2 of the sagging slope, with a higher percentage in obese girls than in normal girls. 2. Serial ECG records during the procedure, performed for all children with the same single-step test, showed differences in the circulatory responses among the test groups. The load was estimated to be 76-86% of the maximal heart rate predicted by Cummings, with obese and girl groups being higher than other groups. The comparison revealed that the heart rate (HR) responses to exercise done by the boys increased faster than those by the girls at 1 minute after exercise. There were differences between obese and control and between boys and girls in HR at recovery, and also in HR before and 4 minutes after exercise between two sexes, which findings agree with other reports. 3. Concerning anthropometric factors, either the skinfold thickness or the obesity index was related to the HR at the peak of the exercise as well as immediately after it and to the half time. By contrast, the arm muscle diameter was found unrelated to the HR. Using this method, applied these combination of Holter ECG and step test to a group examination, we got ECG data for 254 subjects, 97.5% of which were suitable for analysis. We found this method to be effective for detecting ECG changes and for determining HR responses to exercise. This method is safe, simple, and inexpensive and is applicable to examination not only for school children but also for different population groups, for the estimation of circulatory responses.

Age Factors↗