Biomedical subjects

H Mihara

Publications and source records attributed to H Mihara.

At least 55 records · Page 3Linked to original sources

PubMed2000

Gene cloning, purification, and characterization of two cyanobacterial NifS homologs driving iron-sulfur cluster formation.

Iron-sulfur proteins are essential in the photosynthetic system and many other biological processes. We have isolated and characterized enzymes driving the formation of iron-sulfur clusters from Synechocystis sp. PCC6803. Two genes (slr0387 and sll0704), showing similarity to nifS of Azotobacter vinelandii, were cloned, and their gene products (SsCsdl and SsCsd2) were purified. They catalyzed the desulfuration of L-cysteine. Reconstitution of a [2Fe-2S] cluster of cyanobacterial ferredoxin proceeded much faster in the presence of L-cysteine and either of these enzymes than when using sodium sulfide. These results suggest that SsCsdl and SsCsd2 facilitate the iron-sulfur cluster assembly by producing inorganic sulfur from L-cysteine. Synechocystis sp. PCC6803 has no gene coding for a protein with similarity to the N-terminal domain of NifU of A. vinelandii, which is believed to cooperate with NifS to assemble iron-sulfur clusters. Thus, the cluster formation in the cyanobacterium probably proceeds through a mechanism that is different from that in A. vinelandii.

Bacterial Proteins↗

PubMed1999

Design of peptides derived from anti-IgE antibody for allergic treatment.

We have designed and synthesized peptides derived from an anti-IgE antibody which has a potential for the treatment of allergy. It was indicated that conformational restriction of peptide via an intramolecular disulfide bond improved the binding affinity for IgE and that the peptide might have an ability to inhibit the IgE-receptor interaction.

Anti-Allergic Agents↗

PubMed1999

A nifS-like gene, csdB, encodes an Escherichia coli counterpart of mammalian selenocysteine lyase. Gene cloning, purification, characterization and preliminary x-ray crystallographic studies.

Selenocysteine lyase is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the exclusive decomposition of L-selenocysteine to L-alanine and elemental selenium. An open reading frame, named csdB, from Escherichia coli encodes a putative protein that is similar to selenocysteine lyase of pig liver and cysteine desulfurase (NifS) of Azotobacter vinelandii. In this study, the csdB gene was cloned and expressed in E. coli cells. The gene product was a homodimer with the subunit Mr of 44,439, contained 1 mol of PLP as a cofactor per mol of subunit, and catalyzed the release of Se, SO2, and S from L-selenocysteine, L-cysteine sulfinic acid, and L-cysteine, respectively, to yield L-alanine; the reactivity of the substrates decreased in this order. Although the enzyme was not specific for L-selenocysteine, the high specific activity for L-selenocysteine (5.5 units/mg compared with 0.019 units/mg for L-cysteine) supports the view that the enzyme can be regarded as an E. coli counterpart of mammalian selenocysteine lyase. We crystallized CsdB, the csdB gene product, by the hanging drop vapor diffusion method. The crystals were of suitable quality for x-ray crystallography and belonged to the tetragonal space group P43212 with unit cell dimensions of a = b = 128.1 A and c = 137.0 A. Consideration of the Matthews parameter Vm (3.19 A3/Da) accounts for the presence of a single dimer in the crystallographic asymmetric unit. A native diffraction dataset up to 2.8 A resolution was collected. This is the first crystallographic analysis of a protein of NifS/selenocysteine lyase family.

Animals↗

PubMed1999

Optimization of hydrophobic domains in peptides that undergo transformation from alpha-helix to beta-fibril.

Recent studies on peptide fibrillogenesis by the de novo method as well as amyloidogenic proteins including prion proteins and Alzheimer's beta-peptides have provided insights into the conformational changes, such as alpha-helix to beta-structure, involved in folding and misfolding processes. We have found that an exposed hydrophobic nucleation domain at N-terminal causes a structural transition of a peptide from alpha-helix to beta-fibril. It became clear that N-terminal acyl groups of particular lengths in a 2alpha-helix peptide caused the peptide to undergo an alpha-to-beta transition. The peptide with the octanoyl group (C8-2alpha) showed the highest rate of transformation. The study of the designed peptides revealed that these alpha-to-beta transitions were closely related to the initial alpha-helix conformation and its stability. Engineering peptides that undergo alpha-to-beta transitions are attractive not only to the study of pathogenic proteins such as prion proteins, but also to the control of self-assembly of peptides, which will lead to the development of peptidyl self-assembling materials.

Amino Acid Sequence↗

PubMed1999

Design of artificial pepetides that recognize the HIV RRE IIB RNA.

Peptide nucleic acid (PNA) conjugated peptides, derived from HIV-1 Rev, were designed and synthesized in order to construct molecules that recognize HIV RRE IIB RNA. The competitive binding analyses using fluorescent Rev peptide revealed that the PNA unit on the peptide affected the RNA binding.

Amino Acid Sequence↗

PubMed1999

Distribution of JC virus DNA in peripheral blood lymphocytes of hematological disease cases.

OBJECTIVE: The distribution of JC virus DNA in peripheral blood was surveyed by the polymerase chain reaction using the late genes as markers. RESULTS: Six out of 52 cases of hematological diseases and one systemic lupus erythematosus case out of 17 cases were positive for JCV DNA. After separation into B and T lymphocytes by a cell sorter, JCV DNA was found in both cell types prepared from adult T cell leukemia and PML patients. CONCLUSION: Only 1 or 2 copies of JCV genome were calculated to exist in a cell based on the time course analysis of PCR. Only B lymphocytes and glial brain cells are known to produce nuclear factors which support the growth of the virus. The result that B lymphocytes contained a copy number of JCV genome similar to T lymphocytes suggests that there is some barrier to viral growth in susceptible B lymphocytes, and that the growth of JCV is different from that of other virulent viruses.

Aged↗

PubMed1999

[Epstein-Barr virus-associated hemophagocytic syndrome during mid-term pregnancy successfully treated with combined methylprednisolone and intravenous immunoglobulin].

A 32-year-old woman in the 16th week of pregnancy was admitted to our hospital because of high fever. Laboratory findings disclosed pancytopenia and extremely elevated serum LDH and ferritin levels. Coagulation tests showed disseminated intravascular coagulation. Serum soluble interleukin-2 receptor, tumor necrosis factor-alpha, and interleukin-6 levels were high, but serum interferon-gamma was below the detectable limit. Reactive Epstein-Barr virus (EBV) infection was diagnosed on the basis of a high titer of IgG antibodies to the EBV capsid antigen and early antigen. EBV was demonstrated in the peripheral blood and bone marrow cells by polymerase chain reaction. Mature histiocytosis and hemophagocytosis were detected in the bone marrow. A diagnosis of EBV-associated hemophagocytic syndrome (EBV-AHS) was made. Neither prednisolone (PSL 30 mg/day, P.O.) nor methylprednisolone (m-PSL) pulse therapy (1,000 mg/day for 3 days) induced a response. Thereafter, treatment with m-PSL pulse therapy (1,000 mg/day for 3 days) and i.v. administrations of high-dose immunoglobulin (20 g/day for 3 days) in combination with acyclovir (750 mg/day) and gabexate mesilate (2 g/day) induced remission of the disease. Maintenance therapy consisted of PSL (5 mg/day, P.O.) and camostat mesilate (600 mg/day, P.O.). The patient delivered a healthy male infant in the 35th week of pregnancy via natural birth. Reports of pregnant women with EBV-AHS are rare, and the choice of therapy has not yet been established. The present case study suggested the above combination treatment is useful and safe, and capable of changing the fulminant course of EBV-AHS during pregnancy without the use of anticancer drugs.

Acyclovir↗

PubMed1998

Design of novel porphyrin-binding peptides based on antibody CDR.

Novel porphyrin-binding peptides were designed on the basis of an antigen binding site of an antiheme monoclonal antibody. Synthetic peptides were modified with a pyrene moiety. The spectroscopic measurements revealed that the synthetic peptides bound a porphyrin effectively.

Amino Acid Sequence↗

PubMed1998

Design of peptides undergoing self-catalytic alpha-to-beta transition and amyloidogenesis.

Improved understanding of amyloidogenic peptides and proteins such as prion proteins and Alzheimer's beta peptides has attracted much attention to the elucidation of the molecular mechanisms of such amyloidogenesis. As a representative, in the prion protein, the conformational transitions from alpha-helix to beta-structure undergo along with the amyloidogenesis in a self-catalytic manner. Moreover, recent studies by the de novo design of peptides and proteins as well as the amyloidogenesis of peptides and proteins including pathogenic protein mutants have provided insight into the conformational changes essential to amyloidogenesis and correct folding.

Amino Acid Sequence↗

PubMed1998

A method of RF inhomogeneity correction in MR imaging.

A direct postprocessing method for correcting RF inhomogeneity in MR imaging is proposed. First, two images with different flip-angles of theta and 2theta are obtained. Next, the spatial distribution maps of the sensitivity of the surface coil and the B1 field intensity are produced by employing those images. Finally, the correction of the MR image is achieved, dividing the original image by distribution maps of the coil sensitivity and the B1 field intensity. The method was applied to images obtained by a gradient echo sequence and the corrected image is presented.

Humans↗

PubMed1998

Interaction of alpha-helical peptides with phospholipid membrane: effects of chain length and hydrophobicity of peptides.

To investigate the interaction of amphiphilic alpha-helical peptides with phospholipid membranes, we synthesized Ac-(Leu-Ala-Arg-Leu)3-NHCH3 (4[3]) and three derivatives, in which the chain length and the size of the hydrophobic region of the peptides were different from each other. These peptides formed an alpha-helical structure in the presence of vesicles. In the membrane-perturbation measurement, only 43 showed a strong membrane-perturbation activity below phase-transition temperature (25 degrees C), but above phase-transition temperature (50 degrees C), most peptides showed similar strong activities. On the other hand, in membrane-fusion measurement the long peptides, e.g., Ac-(Leu-Ala-Arg-Leu)3-(Leu-Arg-Ala-Leu)3-NHCH3, had strong activities at low peptide concentrations at 25 degrees C. The present study indicated a parallel relationship did not always exist between membrane fusion and perturbation caused by peptides.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1998

Structure and property of model peptides of proline/arginine-rich region in bactenecin 5.

Bactenecin 5 (Bac 5), a cationic antibacterial peptide, contains a repeating region of Arg-Pro-Pro-X (X = hydrophobic residue). To investigate the structure and property of a Pro/Arg-rich region, we synthesized a series of repeating peptides, Ac-(Arg-Pro-Pro-Phe)n-NHCH3 (n = 2, 4, 6, 8 and 10) (PR2, PR4, PR6, PR8 and PR10) as models. The circular dichroism (CD) study suggested that the peptides with longer repeats, PR6, PR8 and PR10, formed a conformation similar to poly(proline)-II in aqueous solution. The CD spectra did not change in the presence of dipalmitoyl-DL-alpha-phosphatidylcholine (DPPC), but they changed in the presence of DPPC/ dipalmitoyl-DL-3-phosphatidylglycerol (DPPG). The gamma-helix, which is very similar in conformation to the poly(proline)-II helix, had the lowest energy conformation for the peptides by energy calculations. Peptides PR6, PR8 and PR10 caused slight leakage of fluorescent dye entrapped in DPPC vesicles, and in the presence of DPPC/DPPG, these peptides showed a considerable level of dye-leakage activity. In contrast, the shorter peptides PR2 and PR4 showed no activity. The same tendency was found in measurements of membrane-fusion activity. Judging from these results, the repeating region of Bac 5 may make a framework to hold a conformation resembling the poly(proline)-II structure in aqueous solution. In addition, this region may interact with acidic lipids, resulting in a change in conformation of the peptide.

Amino Acid Sequence↗

PubMed1998

Use of temporary vena cava filters after catheter-directed fragmentation and thrombolysis in patients with acute pulmonary thromboembolism.

Acute pulmonary thromboembolism is a life-threatening disease and in almost all cases originates from deep venous thrombosis in the proximal deep venous system of the legs. In order to prevent further episodes of thromboembolism in 2 patients with acute pulmonary thromboembolism, we inserted an Antheor temporary vena cava filter after catheter-directed fragmentation and thrombolysis. No complications occurred during the insertion of the filters and our results suggest that this filter device may be clinically safe and effective for temporary protection against pulmonary thromboembolism.

Aged↗

PubMed1997

Cysteine sulfinate desulfinase, a NIFS-like protein of Escherichia coli with selenocysteine lyase and cysteine desulfurase activities. Gene cloning, purification, and characterization of a novel pyridoxal enzyme.

Selenocysteine lyase (EC 4.4.1.16) exclusively decomposes selenocysteine to alanine and elemental selenium, whereas cysteine desulfurase (NIFS protein) of Azotobacter vinelandii acts indiscriminately on both cysteine and selenocysteine to produce elemental sulfur and selenium respectively, and alanine. These proteins exhibit some sequence homology. The Escherichia coli genome contains three genes with sequence homology to nifS. We have cloned the gene mapped at 63.4 min in the chromosome and have expressed, purified to homogeneity, and characterized the gene product. The enzyme comprises two identical subunits with 401 amino acid residues (Mr 43,238) and contains pyridoxal 5'-phosphate as a coenzyme. The enzyme catalyzes the removal of elemental sulfur and selenium atoms from L-cysteine, L-cystine, L-selenocysteine, and L-selenocystine to produce L-alanine. Because L-cysteine sulfinic acid was desulfinated to form L-alanine as the preferred substrate, we have named this new enzyme cysteine sulfinate desulfinase. Mutant enzymes having alanine substituted for each of the four cysteinyl residues (Cys-100, Cys-176, Cys-323, and Cys-358) were all active. Cys-358 corresponds to Cys-325 of A. vinelandii NIFS, which is conserved among all NIFS-like proteins and catalytically essential (Zheng, L., White, R. H., Cash, V. L., and Dean, D. R. (1994) Biochemistry 33, 4714-4720), is not required for cysteine sulfinate desulfinase. Thus, the enzyme is distinct from A. vinelandii NIFS in this respect.

Amino Acid Sequence↗

PubMed1997

Binding of cationic alpha-helical peptides to plasmid DNA and their gene transfer abilities into cells.

Polycationic reagents such as cationic lipids and poly-L-lysine are widely used for gene transfer into cells in vitro and show promise as vectors for in vivo gene therapy applications as nonviral gene transfer techniques. We have developed a novel transfection method using cationic amphiphilic alpha-helical oligopeptides with repeated sequences. Oligopeptide has the advantages of being easily designed and modified because of its simple structure. In this study, we synthesized five kinds of peptides of which the total chain length and the width of the hydrophobic region were changed. The binding of the peptides to plasmid DNA was evaluated by agarose gel electrophoresis. It was found that the long and/or hydrophobic peptides can strongly bind to the DNA. The formation of large aggregates with a 0.5-5-microm diameter, which consisted of the long peptides and the DNA, was observed by electron microscopy. The transfection abilities of the peptides were determined by the expression of luciferase from its cDNA in COS-7 cells. The long peptides showed high transfection abilities. As a result, it could be said that the transfection ability of these peptides was parallel to their ability to form aggregates with DNA. Furthermore, the transfection ability was increased by the addition of chloroquine in the transfection procedure. This result indicated that the internalization of the peptide-DNA aggregates would be mediated by the endocytosis pathway.

Animals↗

PubMed1997

Behavioral correlates of plasma sex hormones and their relationships with plasma lipids and lipoproteins in Japanese men.

In contrast to the hypothesis that endogenous testosterone decreases plasma high-density lipoprotein (HDL) cholesterol levels, many, but not all, studies have reported a positive correlation between plasma total testosterone and HDL cholesterol. We examined behavioral correlates of plasma testosterone and estradiol and the relationships between these sex hormones and plasma lipoproteins, in middle-aged Japanese men. Plasma, lipids, including HDL subfractions, total and free testosterone, and total estradiol were determined with 313 men aged 50-54 years who received a preretirement health examination at the Self-Defence Forces Fukuoka Hospital from January to June in 1992. Body mass index and waist-hip ratio were also measured. Smoking habit, alcohol use, and physical activity were ascertained by a self-administered questionnaire. Obesity, especially waist-hip ratio, was a strong correlate of both total and free testosterone, but not of estradiol. Smoking was associated with elevated levels of testosterone without a dose-effect relation. Neither alcohol use nor physical activity was associated with total or free testosterone, but plasma estradiol levels were higher among current alcohol drinkers. HDL and HDL2 cholesterol were unrelated to either total or free testosterone in the univariate analysis, but negatively associated with free, not total, testosterone after adjustment for obesity. HDL and HDL2 cholesterol also were positively associated with estradiol regardless of adjustment for obesity and other covariates. These findings add to evidence for a hypothesis that high levels of endogenous testosterone and low estradiol levels may cause a decrease in plasma HDL cholesterol, thereby being linked with atherosclerosis in middle aged men.

Alcohol Drinking↗

PubMed1997

Engineering peptides and proteins that undergo alpha-to-beta transitions.

In the 'protein-only' hypothesis, prion diseases are proposed to be the result of conformational changes of the normal form of the prion protein (PrPC) with a highly alpha-helical conformation to a pathogenic scrapie form (PrPSc) with a predominantly beta-sheet conformation. Recent studies examining the folding process of proteins, as well as the amyloidogenesis of peptides and proteins including prion proteins, Alzheimer's beta-peptides and other pathogenic protein mutants, have provided insight into the conformational changes essential to fibrillogenesis and correct folding.

Amino Acid Sequence↗

PubMed1997

Membrane interaction of synthetic peptides related to the putative fusogenic region of PH-30 alpha, a protein in sperm-egg fusion.

In order to investigate the relationship between structure and function of a putative fusogenic region of PH-30a, a protein active in sperm-egg fusion, two peptides, SFP22 and SFP23, whose sequences correspond to the residues 90-111 and 89-111 of PH-30 alpha, respectively, were chemically synthesized. An analog of SFP23, SFP23AA, which has an Ala-Ala sequence instead of the Pro-Pro sequence in SFP23, was also prepared. The CD study indicated that SFP22 and SFP23 mainly took a beta-structure in the presence of DPPC and DPPC/DPPG (3/1) vesicles, while SFP23AA showed an alpha-helical pattern though the alpha-helical content calculated was low (25-30%). alpha-Helical CD curve was observed for these peptides in trifluoroethanol. The membrane-perturbing activity of SFP22 and SFP23 was weaker than that of SFP23AA. On the other hand, the membrane-fusogenic activity of SFP22 and SFP23 to acidic phospholipid bilayers was much stronger than that of SFP23AA. All the peptides caused very weak cell lysis. These results are consistent with the reported speculation [Blobel, C. P. et al. (1992), Nature (London) 356, 248-252] that residues 90-111 of PH-30 alpha may be the fusogenic region and suggest that the Pro-Pro sequence is one of the important factors for holding the active secondary structure of the fusogenic region of PH-30 alpha in membranes.

ADAM Proteins↗