Biomedical subjects

Akira Uchida

Publications and source records attributed to Akira Uchida.

11 recordsLinked to original sources

PubMed2006

Structural mechanism and photoprotective function of water-soluble chlorophyll-binding protein.

A water-soluble chlorophyll-binding protein (WSCP) is the single known instance of a putative chlorophyll (Chl) carrier in green plants. Recently the photoprotective function of WSCP has been demonstrated by EPR measurements; the light-induced singlet-oxygen formation of Chl in the WSCP tetramer is about four times lower than that of unbound Chl. This paper describes the crystal structure of the WSCP-Chl complex purified from leaves of Lepidium virginicum (Virginia pepperweed) to clarify the mechanism of its photoprotective function. The WSCP-Chl complex is a homotetramer comprising four protein chains of 180 amino acids and four Chl molecules. At the center of the complex one hydrophobic cavity is formed in which all of the four Chl molecules are tightly packed and isolated from bulk solvent. With reference to the novel Chl-binding mode, we propose that the photoprotection mechanism may be based on the inhibition of physical contact between the Chl molecules and molecular oxygen.

Binding Sites↗

PubMed2005

The activity of Escherichia coli cyclopropane fatty acid synthase depends on the presence of bicarbonate.

Cyclopropane fatty acid (CFA) synthases catalyze the formation of cyclopropane rings on isolated and unactivated olefinic bonds within various fatty acids; the methylene carbon is derived from the activated methyl group of (S)-adenosylmethionine. The E. coli enzyme is the prototype for this class of enzymes, which include the cyclopropane mycolic acid (CMA) synthases, which are potential targets for the design of antituberculosis agents. Crystal structures of several CMA synthases have recently been solved, and electron density attributed to a bicarbonate ion was found in or near the active site. Because a functional assay for CMA synthases has not been developed, the relevance of the bicarbonate ion has not been established. CFA synthase is 30-35% identical to the CMA synthases that have been analyzed structurally, suggesting that the mechanisms of these enzymes are conserved. In this work, we show that indeed the activity of CFA synthase requires bicarbonate, and that it is inhibited by borate, a planar trigonal molecule that mimics the structure of bicarbonate. We also show that substitutions of the conserved amino acids that act as ligands to the bicarbonate ion based on the structure of CMA synthases result in drastic losses in the activity of the protein.

Bicarbonates↗

PubMed2004

Epimerization of diastereomeric alpha-amino nitriles to single stereoisomers in the solid state.

[reaction: see text] A diastereomeric mixture of the alpha-amino nitrile prepared by the Strecker reaction of benzaldehyde, (1S,2R)-1-aminoindan-2-ol, and cyanotrimethylsilane thermally epimerizes in the solid state to give a single diastereomer with an (S)-configuration at the alpha position to the nitrile moiety. This shows a sharp contrast to the reaction conducted in DMSO at room temperature, which gives a 1:1 mixture of (S)- and (R)-isomers. Several other alpha-amino nitriles also epimerize in the solid-state toward single diastereomers.

Journal Article↗

PubMed2003

Purification, crystallization and preliminary X-ray analysis of a water-soluble chlorophyll protein from Brassica oleracea L. var. acephala (kale).

A water-soluble chlorophyll protein (WSCP) with a chlorophyll a:b ratio of 6:1 from Brassica oleracea L. var. acephala (kale) was purified and crystallized by the hanging-drop vapour-diffusion method using PEG 8000 and zinc acetate as precipitants. The crystal belongs to the hexagonal space group P6(4)22, with unit-cell parameters a = b = 162.2, c = 38.7 A. A native data set was collected to 2.80 A resolution at 293 K using Cu Kalpha radiation from a rotating-anode generator. Preliminary analysis via molecular replacement identified one kale WSCP monomer in the asymmetric unit. The crystal packing showed a tetrameric structure for kale WSCP, as suggested by previous biochemical studies of WSCPs from Brassicaceae plants.

Amino Acid Sequence↗

PubMed2003

Cis,cis,cis-1,2,4,5-cyclohexanetetracarboxylic acid and its dianhydride.

cis,cis,cis-1,2,4,5-Cyclohexanetetracarboxylic acid, C(10)H(12)O(8), (I), contains a mirror plane and the cyclohexane ring exhibits a chair conformation. Two crystallographically independent hydrogen bonds form R(2)(2)(14), R(2)(2)(16) and R(4)(4)(16) ring motifs, and propagation of these two hydrogen bonds along the c and b axes generates C(2)(2)(16) and C(2)(2)(7) chains. cis,cis,cis-1,2:4,5-Cyclohexanetetracarboxylic dianhydride, C(10)H(8)O(6), (II), was prepared by the reaction of (I) with acetic anhydride. The cyclohexane ring of (II) exhibits a boat conformation and the dihedral angle between the two anhydro rings is 117.5 (1) degrees.

Journal Article↗

PubMed2003

Quinolinate dehydrogenase and 6-hydroxyquinolinate decarboxylase involved in the conversion of quinolinic acid to 6-hydroxypicolinic acid by Alcaligenes sp. strain UK21.

In the conversion of quinolinic acid to 6-hydroxypicolinic acid by whole cells of Alcaligenes sp. strain UK21, the enzyme reactions involved in the hydroxylation and decarboxylation of quinolinic acid were examined. Quinolinate dehydrogenase, which catalyzes the first step, the hydroxylation of quinolinic acid, was solubilized from a membrane fraction, partially purified, and characterized. The enzyme catalyzed the incorporation of oxygen atoms of H(2)O into the hydroxyl group. The dehydrogenase hydroxylated quinolinic acid and pyrazine-2,3-dicarboxylic acid to form 6-hydroxyquinolinic acid and 5-hydroxypyrazine-2,3-dicarboxylic acid, respectively. Phenazine methosulfate was the preferred electron acceptor for quinolinate dehydrogenase. 6-Hydroxyquinolinate decarboxylase, catalyzing the nonoxidative decarboxylation of 6-hydroxyquinolinic acid, was purified to homogeneity and characterized. The purified enzyme had a molecular mass of approximately 221 kDa and consisted of six identical subunits. The decarboxylase specifically catalyzed the decarboxylation of 6-hydroxyquinolinic acid to 6-hydroxypicolinic acid, without any co-factors. The N-terminal amino acid sequence was homologous with those of bacterial 4,5-dihydroxyphthalate decarboxylases.

Alcaligenes↗

PubMed2002

Difference between left and right lateral ventricular sizes in neonates.

The objective of this study is to determine the causes of asymmetry of the lateral ventricles in neonates. We also studied the effect of head position and the relationship of body weight at birth in regard to lateral ventricular size. Eligible for inclusion in this study were 60 neonatal infants whose gestational age was 33.1+/-3.5 weeks and whose birth weight was 1793+/-613 g. Ultrasonographic examinations were performed at the first and the second weeks after birth. In parasagittal and coronal scans through the posterior horn of the lateral ventricle, the lateral ventricle was traced and its area was measured. We found no significant variation of ventricular size in relation to body weight at birth. The left ventricular size was larger than the right one. The difference of the left and right ventricular sizes was partially effected by head position. The ratio of left to right lateral ventricular sizes showed a very wide distribution. We considered that ventricular asymmetry is not pathological, but due to individual differences.

Aging↗

PubMed2002

Effect of head position to the cerebral arterial flow in neonates.

The objective of this study is to determine the difference of the flow velocities of left and right cerebral arteries. We also studied the effect of head position to the cerebral arterial flow velocities. Eligible for inclusion in this study were 60 neonatal infants whose gestational age was 33.1+/-3.5 weeks and whose birth weight was 1793+/-613 g. The ultrasonographic examinations were performed in the first and second weeks after birth. In an axial scan through a temporal window, the Doppler sample volume was positioned at the center of the M1 portion of the middle cerebral artery and the flow velocity curve was detected. No statistical difference was seen in the flow velocities between the left and right middle cerebral arteries. However, the flow velocities in the upper side were significantly higher than those in the lower side. RI in the upper side was significantly smaller than that in the lower one. This change of flow velocities stabilized in 5 min after the head was turned upside down. The effect of head positioning to the intracranial blood flow must be considered when cerebral ultrasonography of neonates is performed.

Blood Flow Velocity↗

PubMed2002

Magnetoencephalographic study of speed-dependent responses in apparent motion.

OBJECTIVES: There have been only few studies of visually-evoked cortical responses to apparent motion as a function of stimulus speed. Most earlier findings on evoked peak magnitudes and latencies, utilizing various types of smooth and apparent motion stimuli, have demonstrated that greater spatial separation/speed resulted in enhanced peak magnitudes, decreasing onset latencies in individual extrastriate neurons and in shorter motor reaction times in subjects. However, some reports using partial-coverage magnetoencephalography stated that increasing the stimulus displacement actually triggered a substantial reduction of the evoked main peak latency while the magnitude showed no clear change. METHODS: To resolve the issue of the dependency of evoked responses on stimulus speed in apparent motion, we presented moving bar stimuli to 6 subjects at velocities within a 100 fold range and investigated the ensuing evoked visual cortical activity using a whole-cortex magnetoencephalograph. The magnitude and the latency of the first major evoked peak M1 was measured and compared for 6 discrete bar-stimuli displacements in all subjects. RESULTS: Our results showed clearly that the M1 peak response magnitudes increased in a nonlinear way with higher apparent speeds (larger displacements), in compliance with the logarithmic Fechner law. We observed also that the fluctuations of the mean evoked M1 peak latency (140+/-10.6 ms) did not reach significance over the tested range of stimulus velocities. CONCLUSIONS: These findings probably reflect global motion processing mechanisms which rely on nonlinear speed-dependent feedback connectivity between striate and extrastriate visual cortex areas.

Adult↗

PubMed2002

The influence of Ala243 (Gly247), Arg215 and Thr226 (Asn230) on the bioluminescence spectra and pH-sensitivity of railroad worm, click beetle and firefly luciferases.

Among beetle luciferases, the pH-sensitive firefly luciferases have been studied extensively. Much less is known about pH-insensitive luciferases, which include click beetle and railroad worm luciferases. Previously, we found that the residues R215 and T226 (N230) are important for green light emission. Here we show that the conserved residue A243 in pH-insensitive luciferases and the corresponding G247 in pH-sensitive luciferases affect the emission spectrum and influence pH-sensitivity. In contrast to railroad worm green light-emitting (PxvGR) and firefly luciferases, the substitution of R215 in Pyrearinus termitilluminans click beetle luciferase (Pte) had no effect on the spectrum, showing that R215 is not essential for green light emission in all beetle luciferases. A homology-based model of Pte luciferase shows that R215 and T226 are close enough to interact. To investigate if there was an interaction between these conserved residues, double mutants were constructed. The double substitution R215S/T226N in Pte luciferase abolished the activity. In PxvGR luciferase the same double mutant resulted in a redshift (lambda(max) = 595 nm), whose magnitude was lower than the value expected for an additive effect. These results suggest that the effects of R215S and T226N are partially interdependent. The double substitution T226N/A243G had an additive redshift effect on the spectrum of PxvGR luciferase, whereas it had a smaller effect on the spectrum of Pte luciferase. Altogether, these results suggest that the above substitutions have different effects on the active site of click beetle and railroad worm luciferases.

Alanine↗

PubMed1999

High-Spin (meso-Tetraalkylporphyrinato)iron(III) Complexes As Studied by X-ray Crystallography, EPR, and Dynamic NMR Spectroscopies.

1H NMR spectra of a series of high-spin (meso-tetraalkylporphyrinato)iron(III) chlorides, [Fe(TRP)Cl] where R = Me, Et, Pr, or (i)Pr, have been measured at various temperatures in CD(2)Cl(2) solution. In the case of the Et, Pr, and (i)Pr complexes, either the methyl or the methylene signal split into two signals with equal integral intensities at low temperature. In contrast, the Me complex did not show any splitting even at -100 degrees C. The results have been ascribed to the hindered rotation of the meso-alkyl groups about C(meso)-C(alpha) bonds. The activation free energies for rotation have been determined as 8.0 (-72 degrees C), 8.5 (-60 degrees C), and 8.9 (-62 degrees C) kcal.mol(-1) for the Et, Pr, and (i)Pr complexes, respectively, at coalescence temperatures given in parentheses. The small activation free energy for rotation of the isopropyl groups observed in the present system is explained in terms of the nonplanarity of the porphyrin ring, which has been verified both by the X-ray crystallographic analysis and by the EPR spectrum taken in a frozen CH(2)Cl(2)-toluene solution. The success in observing the hindered rotation of less bulky primary alkyl groups such as ethyl and propyl groups at an easily accessible temperature range is attributed to the large difference in chemical shifts of the mutually exchanging protons, ca. 3500 Hz in the case of the Et complex, caused by the paramagnetism of the five-coordinated ferric porphyrin complexes.

Journal Article↗