Publications by authors named "Kiyohiro Imai"

Human hemoglobin (Hb), which is an α2β2 tetramer and binds four O2 molecules, changes its O2-affinity from low to high as an increase of bound O2, that is characterized by 'cooperativity'. This property is indispensable for its function of O2 transfer from a lung to tissues and is accounted for in terms of T/R quaternary structure change, assuming the presence of a strain on the Fe-histidine (His) bond in the T state caused by the formation of hydrogen bonds at the subunit interfaces. However, the difference between the α and β subunits has been neglected.

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Incorporation of the heme into globin induces a prominent circular dichroism (CD) band in the Soret region. The appearance of heme optical activity is widely believed to arise from the interaction between the heme and aromatic residues of the globin. However, hemoglobin (Hb) containing the reversed heme exhibits a CD spectrum obviously different from that of native Hb, indicating that the interactions of heme side chains with globin contribute to the appearance of heme optical activity.

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The quaternary structures of invertebrate haemoglobins (Hbs) are quite different from those of vertebrate Hbs. The extracellular giant Hbs of molecular masses of about 400 and 3600 kDa are composed of a dome-shaped dodecameric subassembly which consists of four individual globin subunits. Several crystal structures of 400 kDa Hbs from annelids have been reported, including structures in oxygenated and partially unliganded states, but the structure of the fully deoxygenated state has not been reported.

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While heme alone does not exhibit circular dichroism (CD) spectrum, it exhibits a prominent positive CD band in the Soret region when incorporated into apoglobin of myoglobin (Mb) and hemoglobin (Hb). The appearance of this optical activity is widely accepted to arise from the interactions between the heme and aromatic residues of the globin. However, the reversed heme orientation in Hb was found to exhibit a CD spectrum obviously different from that of native Hb, indicating that the interactions of side chains of heme with globin also contribute to the appearance of heme optical activity.

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We analyzed the oxygen (O2) and carbon monoxide (CO) binding properties of the H64L mutant of myoglobin reconstituted with chemically modified heme cofactors possessing a heme Fe atom with a variety of electron densities, in order to elucidate the effect of the removal of the distal His64 on the control of both the O2 affinity and discrimination between O2 and CO of the protein by the intrinsic heme Fe reactivity through the electron density of the heme Fe atom (ρFe). The study revealed that, as in the case of the native protein, the O2 affinity of the H64L mutant protein is regulated by the ρFe value in such a manner that the O2 affinity of the protein decreases, due to an increase in the O2 dissociation rate constant, with a decrease in the ρFe value, and that the O2 affinities of the mutant and native proteins are affected comparably by a given change in the ρFe value. On the other hand, the CO affinity of the H64L mutant protein was found to increase, due to a decrease in the CO dissociation rate constant, with a decrease in the ρFe value, whereas that of the native protein was essentially independent of a change in the ρFe value.

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We analyzed the vibrational frequencies of the Fe-bound carbon monoxide (CO) of myoglobin reconstituted with a series of chemically modified heme cofactors possessing a heme Fe atom with a variety of electron densities. The study revealed that the stretching frequency of Fe-bound CO (ν(CO)) increases with decreasing electron density of the heme Fe atom (ρ(Fe)). This finding demonstrated that the ν(CO) value can be used as a sensitive measure of the ρ(Fe) value and that the π back-donation of the heme Fe atom to CO is affected by the heme π-system perturbation induced through peripheral side chain modifications.

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Studies using myoglobins reconstituted with a variety of chemically modified heme cofactors revealed that the oxygen affinity and autoxidation reaction rate of the proteins are highly correlated to each other, both decreasing with decreasing the electron density of the heme iron atom. An Fe(3+)-O(2)(-)-like species has been expected for the Fe(2+)-O(2) bond in the protein, and the electron density of the heme iron atom influences the resonance process between the two forms. A shift of the resonance toward the Fe(2+)-O(2) form results in lowering of the O(2) affinity due to an increase in the O(2) dissociation rate.

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Myoglobin (Mb) stores dioxygen in muscles, and is a fundamental model protein widely used in molecular design. The presence of dimeric Mb has been known for more than forty years, but its structural and oxygen binding properties remain unknown. From an X-ray crystallographic analysis at 1.

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The aromatic residues such as tryptophan (Trp) and tyrosine (Tyr) in human adult hemoglobin (Hb A) are known to contribute to near-UV circular dichroism (CD) and UV resonance Raman (RR) spectral changes upon the R → T quaternary structure transition. In Hb A, there are three Trp residues per αβ dimer: at α14, β15, and β37. To evaluate their individual contributions to the R → T spectral changes, we produced three mutant hemoglobins in E.

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To test liposome-encapsulated hemoglobin (LEH) in transient cochlear ischemia/reperfusion as a model of sudden deafness, Mongolian gerbils were randomly assigned to receive 2 mL/kg of either low-affinity LEH (l-LEH, P₅₀0₂ = 40 mm Hg), high-affinity LEH (h-LEH, P₅₀0₂ = 10 mm Hg), homologous red blood cells (RBCs), or saline (each group n = 6) 30 min before 15-min occlusion of the bilateral vertebral arteries and reperfusion. Sequential changes in hearing were assessed by auditory brain response 1, 4, and 7 days after ischemia/reperfusion, when the animals were sacrificed for pathological studies. h-LEH was significantly more protective than l-LEH in suppressing hearing loss, in contrast to RBC or saline treatment, at 8, 16, and 32 kHz, where hearing loss was most severe (P < 0.

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A pair of myoglobins containing inherently distorted alpha-ethyl-2,4-dimethyldeuteroheme or undistorted 2,4-dimethyldeuteroheme were prepared, and the functional consequence of intrinsic heme deformation was investigated. The visible absorption peaks of the myoglobin bearing the distorted heme exhibited a bathochromic shift, indicating that the heme was deformed in the protein pocket. Ligand affinities for the ferric myoglobin with the distorted heme were found to be higher than those of the myoglobin bearing the undistorted heme.

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Functional regulation of myoglobin (Mb) is thought to be achieved through the heme environment furnished by nearby amino acid residues, and subtle tuning of the intrinsic heme Fe reactivity. We have performed substitution of strongly electron-withdrawing perfluoromethyl (CF(3)) group(s) as heme side chain(s) of Mb to obtain large alterations of the heme electronic structure in order to elucidate the relationship between the O(2) affinity of Mb and the electronic properties of heme peripheral side chains. We have utilized the equilibrium constant (pK(a)) of the "acid-alkaline transition" in metmyoglobin in order to quantitatively assess the effects of the CF(3) substitutions for the electron density of heme Fe atom (rho(Fe)) of the protein.

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Among the four types of hemoglobin (Hb) M with a substitution of a tyrosine (Tyr) for either the proximal (F8) or distal (E7) histidine in the alpha or beta subunits, only Hb M Saskatoon (betaE7Tyr) assumes a hexacoordinate structure and its abnormal subunits can be reduced readily by methemoglobin (metHb) reductase. This is distinct from the other three M Hbs. To gain new insight into the cause of the difference, we examined the ionization states of E7 and F8 Tyrs by UV resonance Raman (RR) spectroscopy and Fe-O(Tyr) bonding by visible RR spectroscopy.

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Fatty acid transport protein (FATP) is an evolutionarily conserved membrane-bound protein that facilitates the uptake of extracellular long chain fatty acids. In humans and mice, six FATP isoforms have been identified and their tissue-specific distributions suggest that each plays a discrete role in lipid metabolism in association with fatty acid uptake. While the presence of FATP homologs in insects has been demonstrated, their functional role remains to be characterized.

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The iron complex of oxypyriporphyrin, a porphyrinoid containing a keto-substituted pyridine, was coupled with apomyoglobin. The reconstituted ferric myoglobin was found to be five-coordinate without iron-bound water molecules. The anionic ligands such as CN (-) and N 3 (-) bound the myoglobin with high affinities, while neutral imidazole did not.

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Objectives: This study examined whether a reduction in hemoglobin-oxygen affinity improves exercise capacity in mice with heart failure.

Background: Exercise intolerance is a major determinant of quality of life in patients with chronic heart failure. One of the major goals of the treatment for chronic heart failure is to improve quality of life.

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We found that recombinant human adult hemoglobin (rHb A) expressed in Escherichia coli showed heterogeneity of components with the intensity of a positive CD band at 260 nm and that it could be resolved into three components (SP-1, SP-2, and SP-3) by SP-Sepharose column chromatography. 1H NMR revealed that SP-1 is identical with native Hb A, while SP-2 and SP-3 largely contain the reversed heme isomer in both the alpha and beta subunits, with contents of approximately 50 and >80% in SP-2 and SP-3, respectively. Rotation of the heme 180 degrees about the 5,15-meso axis (reversed heme) causes an interexchange of the methyl groups at positions 2 and 7 with the vinyl groups at positions 8 and 3, respectively.

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In most female moths, pheromone biosynthesis activating neuropeptide (PBAN) regulates sex pheromone production by stimulating an influx of extracellular Ca(2+). Little is known about the plasma membrane channel or how the PBAN stimulus is communicated to the channel. Fluorescent Ca(2+) imaging techniques confirmed PBAN-induced Ca(2+) influx in the silkworm, Bombyx mori, and showed that the PBAN response is reduced with repeated stimulation.

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Oxygenation properties of hemoglobin (Hb) from Oligobrachia mashikoi were extensively investigated. Compared to human Hb, Oligobrachia Hb showed a high oxygen affinity (P(50)=1.4 mmHg), low cooperativity (n =1.

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In most moths, sex pheromone production is regulated by pheromone biosynthesis-activating neuropeptide (PBAN). How the extracellular PBAN signal is turned into a biological response has been the focus of numerous studies. In the classical scheme of signal transduction, activated G proteins relay the extracellular signal to downstream effector molecules such as calcium channels and adenylyl cyclase.

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A ring fluorinated heme, 13,17-bis(2-carboxylatoethyl)-3,8-diethyl-2-fluoro-7,12,18-trimethyl-porphyrinatoiron(III), has been incorporated into human adult hemoglobin (Hb A). The heme orientational disorder in the individual subunits of the protein has been readily characterized using (19)F NMR and the O(2) binding properties of the protein have been evaluated through the oxygen equilibrium analysis. The equilibrated orientations of hemes in alpha- and beta- subunits of the reconstituted protein were found to be almost completely opposite to each other, and hence were largely different from those of the native and the previously reported reconstituted proteins [T.

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The O2 binding properties of bovine Hb were examined. The increase in Cl- and DPG concentration enhanced P50. A reduction in n(max) was observed at high Cl- concentration, while DPG had little effect on n(max).

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The iron complex of a new type of corrphycene bearing two ethoxycarbonyl (-CO2C2H5) groups on the bipyrrole moiety was introduced into apomyoglobin. The reconstituted ferric myoglobin has a coordinating water molecule that deprotonates to hydroxide with a pK(a) value of 7.3 and exhibits 3-10-fold higher affinities for anionic ligands when compared with a counterpart myoglobin with the same substituents on the dipyrroethene moiety.

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By using published experimental values of the standard oxygen (O2) equilibrium curve and the in vivo arterial and venous O2 pressure (PO2) of fetal and maternal blood in five mammalian species (human, cow, pig, sheep, and horse), we investigated the relationship between the efficiency of O2 delivery and the effectiveness of the Bohr shift, and discussed the significance of cooperativity for mammalian Hb. The O2 delivery of fetal blood was more efficient than that of maternal blood, and the effectiveness of the Bohr shift at both O2 loading and release sites of fetal blood was high. A linear relationship was observed between the efficiency of O2 delivery and the effectiveness of the Bohr shift at O2 loading sites of the five mammalian species.

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A novel hemoglobin (Hb) variant was found in a specimen that showed an unusual profile in analyses of glycohemoglobin An abnormal beta-globin, 443 Da smaller than normal beta-globin, was detected by electrospray ionization mass spectrometry (ESI/MS) with intact globin. Mass spectrometry analysis of tryptic peptides derived from isolated abnormal Hb showed an abnormal peptide, characterized as betaT- 14 (141Leu-->Val and 144Lys-->0). Nucleotide sequencing revealed a heterozygosity of codon 141 CTG(Leu)-->GTG(Val), and codon 144 AAG(Lys)-->TAG(stop codon).

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