Phenomenology of superstring-inspired SO(10) grand-unified-theory model with intermediate scale.
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Journal Article↗
Biomedical subjects
H Okada
Publications and source records attributed to H Okada.
Polymerization of factor B of the alternative complement pathway via disulfide bond(s) in the presence of Cu2+ and stimulation by C3b, the major fragment of C3.
Factor B(B) of the alternative complement pathway has been found to dimerize via disulfide bond(s) in the presence of CuCl2. Poly B has no B activity. The Bb fragment was also dimerized, indicating that one free sulfhydryl group on the Bb portion might be involved in polymerization. The Ba fragment was not dimerized. C3b, the major fragment of C3, has the capacity to stimulate polymerization of B. Incubation of C3b, B and factor D in the presence of Mg2+ and Cu2+ resulted in the formation of poly B and diminished cleavage of B. These results suggest that polymerization of B due to Cu2+ might be partly responsible for the impairment of C3 convertase activity of the alternative pathway.
Complement C3↗
Determination of the active-site serine of 6-aminohexanoate-dimer hydrolase.
Diisopropylfluorophosphate, an inhibitor of serine proteinase, was used to label 6-aminohexanoate-dimer hydrolase, a nylon oligomer degradative enzyme of Flavobacterium sp. K172. More than 95% of the enzyme activity was lost upon incorporation of 1-1.5 molecules inhibitor/subunit of the enzyme. The tryptic peptide of the labeled enzyme was purified by HPLC (reverse-phase partition) and its amino acid sequence was identified. Radioactivity was found to be incorporated into an 8-amino-acid peptide (108His-Leu-Leu-Met-Ser-Val-Ser-Lys115). Amino acid alteration from Ser to Ala at the position 112 by site-directed mutagenesis caused loss of enzyme activity to below the detection threshold (1% of the activity of the parental enzyme). These results indicate that Ser112 is essential for the activity.
Affinity Labels↗
Complete deficiency of 20 KDa homologous restriction factor (HRF20) and restoration with purified HRF20.
20 KDa homologous restriction factor (HRF20) is a membrane glycoprotein which inhibits formation of membrane attack complexes of homologous complement. Erythrocytes from a patient who is completely deficient in HRF20 were readily hemolyzed by homologous complement activated by sucrose or by acidification as in paroxysmal nocturnal hemoglobinuria (PNH). After incubating PNH erythrocytes (PNH-E) with purified HRF20, the cells were analyzed by flow cytometry using a monoclonal antibody to HRF20 and shown to have the antigen absorbed. These PNH-E acquired resistance to hemolysis by homologous complement suggesting that HRF20 may be successfully used for treatment of these patients.
Adult↗
Monoclonal antibodies capable of causing hemolysis of neuraminidase-treated human erythrocytes by homologous complement.
As human E (HuE) treated with neuraminidase (Neu) are resistant to hemolysis by human serum but are readily lysed by heterologous serum via the alternative C pathway, we attempted to produce mAb which might modify Neu-treated HuE (Neu-HuE) so as to render them sensitive to homologous C. A hybridoma, clone -1F5, was obtained by screening for antibody which caused hemolysis of Neu-HuE by human serum via the alternative C pathway. We have shown that this antibody (1F5) of IgG1 isotype blocks the action of a 20-kDa membrane inhibitor capable of interfering with the terminal step in the homologous C cascade. The antigenic molecule can be termed HRF20, which stands for homologous restriction factor (HRF) with m.w. 20,000, because its function is essentially the same as that of HRF (68,000) reported by others.
Animals↗
Assay of complement activity in human serum using large unilamellar liposomes.
Liposomes incorporating 2,4,6-trinitrophenyl-aminocaproyl-dipalmitoylphosphatidylethanolamine (TNP-cap-DPPE) as a membrane hapten can be lysed by human complement in the presence of anti-TNP antibody. Liposomes were composed of L-alpha-dimyristoylphosphatidylcholine, cholesterol, TNP-cap-DPPE and dicetylphosphate at a molar ratio of 1:1:0.005:0.02. Large unilamellar liposomes were prepared by reverse-phase evaporation with an entrapped marker, carboxyfluorescein, which is self-quenching in liposomes at high concentrations such as 0.2 M. When the marker is released by complement action, a strong fluorescence is produced. The amount of marker released from liposomes is able to quantify complement activity of both the classical and alternative pathways. CL50 is the complement activity obtained by lysis of 50% of the liposomes. The number of CL50 units in sera obtained with liposomes correlates well with those obtained using the hemolytic complement test CH50 (r = 0.98). The advantages of this method include stability of reagents, accuracy, simplicity and speed. In addition, it is well suited for developing an automated system. This method is a useful substitute for the hemolytic assay for determination of human complement activity.
Complement System Proteins↗
An arginine specific carboxypeptidase generated in blood during coagulation or inflammation which is unrelated to carboxypeptidase N or its subunits.
An unstable carboxypeptidase N or B like enzyme is generated as a result of coagulation. This enzyme is derived from some plasma component (s) and not from blood cells or platelets. Furthermore, the activity generated is specific for arginine substrates insofar as small synthetic substrates are concerned. The enzyme is unrelated to CPN or any of its subunits or subunit fragments. This transient carboxypeptidase may be involved in the processing and/or scheduling of different functions of bioactive peptides generated during inflammation.
Arginine↗
20 KDa homologous restriction factor of complement resembles T cell activating protein.
We previously identified a 20KDa membrane glycoprotein 1F5 antigen which inhibits the assembly of homologous complement membrane attack complexes and we designate it as HRF20 standing for 20KDa homologous restriction factor. The amino acid sequence deduced from its coding base sequence resembles that of T cell activating protein, most conspicuously in cysteine residues, 10 out of 11 of which occupy identical positions in an overall sequence homology of 24.8%. Furthermore, proliferation of human T cells was stimulated by monoclonal antibody to HRF20.
Amino Acid Sequence↗
Stability-increasing mutants of glucose dehydrogenase.
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Bacillus megaterium↗
Enzymatic method for measuring the absolute value of oxygen concentration.
An enzymatic method for measuring the absolute concentration of oxygen in aqueous solutions, using 4-hydroxybenzoate 3-monooxygenase and glucose oxidase, is described. The monooxygenase is used for quantitative oxidation of 4-hydroxybenzoate and NADPH with oxygen into 3,4-dihydroxybenzoate and NADP+; the amount of oxygen can be measured as the amount of NADPH decreased by the reaction. The monooxygenase reaction is performed in a syringe, a closed system. After the completion of the monooxygenase reaction, glucose oxidase is added to the assay solution to consume the oxygen from the atmosphere; this enables us to measure the NADPH concentration in the solution spectrophotometrically in an open system and to check the anaerobicity of closed systems. The oxygen concentrations at 25 degrees C of oxygen-saturated and air-saturated water were 1.10 and 0.23 mM, respectively. The value for argon-bubbled water was zero within the experimental error; this justifies the assay system. Thus, it is shown that a sample containing 8 microM-1.1 mM oxygen can be measured by this method.
Aspergillus↗
Stability-increasing mutants of glucose dehydrogenase from Bacillus megaterium IWG3.
A glucose dehydrogenase gene was isolated from Bacillus megaterium IWG3, and its nucleotide sequence was identified. The amino acid sequence of the enzyme deduced from the nucleotide sequence is very similar to the protein sequence of the enzyme from B. megaterium M1286 reported by Jany et al. (Jany, K.-D., Ulmer, W., Froschle, M., and Pfleiderer, G. (1984) FEBS Lett. 165, 6-10). The isolated gene was mutagenized with hydrazine, formic acid, or sodium nitrite, and 12 clones (H35, H39, F18, F20, F191, F192, N1, N13, N14, N28, N71, and N72) containing mutant genes for thermostable glucose dehydrogenase were obtained. The nucleotide sequences of the 12 genes show that they include 8 kinds of mutants having the following amino acid substitutions: H35 and H39, Glu-96 to Gly; F18 and F191, Glu-96 to Ala; F20, Gln-252 to Leu; F192, Gln-252 to Leu and Ala-258 to Gly; N1, Glu-96 to Lys and Val-183 to Ile; N13 and N14, Glu-96 to Lys, Val-112 to Ala, Glu-133 to Lys, and Tyr-217 to His; N28, Glu-96 to Lys, Asp-108 to Asn, Pro-194 to Gln, and Glu-210 to Lys; and N71 and N72, Tyr-253 to Cys. These mutant enzymes have higher stability at 60 degrees C than the wild-type enzyme. The results of this study indicate that the tetrameric structure of glucose dehydrogenase is stabilized by several kinds of mutation, and at least one of the following amino acid substitutions stabilizes the enzyme: Glu-96 to Gly, Glu-96 to Ala, Gln-252 to Leu, and Tyr-253 to Cys.
Amino Acid Sequence↗
Structure and properties of malic enzyme from Bacillus stearothermophilus.
The malic enzyme (EC 1.1.1.38) gene of Bacillus stearothermophilus was cloned in Escherichia coli, and the enzyme was purified to homogeneity from the E. coli clone. In addition to the NAD(P)-dependent oxidative decarboxylation of L-malate, the enzyme catalyzes the decarboxylation of oxalacetate. The enzyme is a tetramer of Mr 200,000 consisting of four identical subunits of Mr 50,000. The pH optima for malate oxidation and pyruvate reduction are 8.0 and 6.0, respectively; and the optimum temperature is 55 degrees C. The enzyme strictly requires divalent metal cations for its activity, and the activity is enhanced 5-7 times by NH4+ and K+. Kinetic study shows that the values of the dissociation constant of the enzyme-coenzyme complex are 77 microM for NAD and 1.0 mM for NADP, indicating that the enzyme has a higher affinity for NAD than for NADP. The nucleotide sequence of the gene and its flanking regions was also found. A single open reading frame of 1434 base pairs encoding 478 amino acids was concluded to be that for the malic enzyme gene because the amino acid composition of the enzyme and the sequence of 16 amino acids from the amino terminus of the enzyme agreed well with those deduced from this open reading frame.
Amino Acid Sequence↗
Synthesis and characterization of 1-substituted 5-alkylphenazine derivatives carrying functional groups.
The following 1-substituted derivatives of 5-methylphenazine and 5-ethylphenazine were synthesized: 1-(3-carboxypropyloxy)-5-methylphenazine (1B), 1-(3-carboxypropyloxy)-5-ethylphenazine (2B), 1-(3-ethoxycarbonylpropyloxy)-5-ethylphenazine (2C) and 1-[N-(2-aminoethyl)carbamoylpropyloxy]-5-ethylphenazine (2D); their spectra, stability and reactivity as electron mediators were investigated, together with those of 5-methylphenazine (1A) and 5-ethylphenazine (2A). The 1-substituted derivatives are all insensitive to light and the derivatives of 5-ethylphenazine are more stable than those of 5-methylphenazine under neutral and alkaline conditions; 2B is the most stable of all the derivatives. The spectral properties of the decomposed compounds showed that photodecomposition of 1A and 2A is associated with hydroxylation at position 1, alkali decomposition of 1A and 1B with elimination of the 5-methyl group and alkali decomposition of 2A, 2B, and 2D with a ring-opening reaction. The second-order rate constant k1 for the reaction of the phenazine derivatives with NADH was measured under steady-state conditions. The k1 values vary depending on the substituents at positions 1 and 5: the values for 1A, 1B, 2A, 2B, 2C and 2D are 1.83 mM-1 s-1, 3.33 mM-1 s-1, 0.75 mM-1 s-1, 1.42 mM-1 s-1, 1.68 mM-1 s-1 and 2.03 mM-1 s-1, respectively. The rate constants, k2 and k3, for the reactions of the reduced form of 2B with oxygen and with 3-(4',5'-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium ion, respectively, were k2 = 1.21 mM-1 s-1 and k3 = 91 mM-1 s-1. These phenazine derivatives have potential applications in the biochemical field.
Alkylation↗
Preparation and kinetic properties of 5-ethylphenazine-poly(ethylene glycol)-NAD+ conjugate, a unique catalyst having an intramolecular reaction step.
5-Ethylphenazine-poly(ethylene glycol)-NAD+ conjugate (EP+-PEG-NAD+) was prepared by linking 1-(3-carboxypropyloxy)-5-ethylphenazine (I) to poly(ethylene glycol)-bound NAD+ (PEG-NAD+) and its kinetic properties were studied. As a reference compound, poly(ethylene glycol)-bound 5-ethylphenazine derivative (III) was also prepared and the effects of poly(ethylene glycol) on the reaction rate of the 5-ethylphenazine moiety with NADH was investigated. The second-order rate constant, k1, of the reaction of III with NADH is 2.78 mM-1 s-1 and is about 1.7 times that of 1-(3-ethoxycarbonylpropyloxy)-5-ethylphenazine (II) with NADH. A similar effect of the attached poly(ethylene glycol) was observed for the reaction of PEG-NADH with I or II. The second-order rate constants, k2 and k3, of the reactions of the reduced form of III with oxygen and with 3-(4',5'-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium ion, respectively, were k2 = 1.22 mM-1 s-1 and k3 = 32 mM-1 s-1; the k2 value is not changed but the k3 value is decreased by the attachment of the polymer. EP+-PEG-NAD+ works as a unique catalyst having an intramolecular reaction step within its turnover cycle in a coupled multi-step reaction system containing malate dehydrogenase, malate, EP+-PEG-NAD+, a tetrazolium salt and oxygen. The first-order rate constant, k4, of the intramolecular reaction was 1.1 s-1. The effects of the covalent linking of the 5-ethylphenazine and the NAD+ moieties were estimated by comparing the value of k4 with that of k1 for the reaction of III with NADH; the effective concentration of the NADH moiety for the 5-ethylphenazine moiety on the same EP+-PEG-NADH molecule (or vice versa) was calculated to be 0.40 mM from the ratio of k4/k1. The values of the rate constants in the coupled multi-step reaction system enable us to understand the dynamic features of the system and the characteristics of EP+-PEG-NAD+ as a catalyst are discussed.
Catalysis↗
Prognosis of cervical cancers with reference to steroid receptors.
The relationship among the prognosis of a patient with cancer of the cervix and endocrine background [production of estrogen receptor (ER), progestogen receptor (PR), and androgen receptor (AR)], clinical stage, histological type, lymph node metastasis, age, and treatment was studied. ERs were detected in almost all cases, regardless of the histological type of cancer. The detection rate and level of PR were in the decreasing order of large-cell non-keratinizing (LNK) carcinoma, keratinizing (K) carcinoma, and small-cell non-keratinizing (SNK) carcinoma of the cervix. Thus, the presence and level of PR in squamous cell carcinoma of the cervix were considered to be an indicator of the degree of differentiation. PR was not detected in cases of adenocarcinoma of the cervix. This may explain the poor responsiveness to progestogen treatment in adenocarcinoma of the cervix. There was no relationship between the patient's prognosis and the presence of PR or AR, but the prognosis of patients with lesions rich in ER (greater than or equal to 5 fmol/micrograms DNA) tended to be better than that of patients with lesions poor in ER (less than 5 fmol/micrograms DNA). The prognosis of patients before menopause tended to be better than that postmenopause. These findings indicate a relationship between cancer of the cervix and endocrine background.
Adenocarcinoma↗
The inductive effects of progestogens on aromatase activity in stromal cells of human uterine endometrium.
Aromatase activity for androstenedione was measured in stromal cells of human uterine endometrium throughout the menstrual cycle. The effect of progestogen on aromatase activity was also examined in the endometrial stromal cells. Aromatase activity tended to be high in the proliferative cells compared to the secretory cells. Moreover, its enzyme activity was enhanced at the concentration of 0.1 or 1.0 microM of medroxyprogesterone acetate (MPA) and norethindrone (ENT) in all phases of menstrual cycle. MPA stimulated aromatase activity in the stromal cells more strongly than did ENT. These results may suggest that although aromatase in uterine endometrium is not regarded to be so much influenced by endogenous progesterone during menstruation, the increase of aromatase activity by progestogen added lead to the more differentiated endometrium.
Aromatase↗
Effects of stimulation of the hypothalamic area on pancreatic exocrine secretion in dogs.
The effects of electrical or chemical (0.1 M dl-homocysteic acid) stimulation of the hypothalamus on pancreatic exocrine secretion were studied in chloralose-anesthetized and hemispherectomized dogs whose pyloric sphincter had been ligated. Excitatory pancreatic flow responses with frequently increased antral contractility and small changes in blood pressure were induced by stimulation of the ventral and dorsal portions of the anterior hypothalamic area, the lateral part of the middle hypothalamus, and the mamillary body. The inhibitory pancreatic responses with reduced antral and corpus contractility and elevated blood pressure were elicited by stimulation of the posterior hypothalamic area, the middle portion of the anterior hypothalamus and the most dorsal area of the hypothalamus. Both excitatory and inhibitory responses were obtained even in dogs with cervical cord transection. The excitatory responses and some of the inhibitory ones were abolished by vagotomy or atropine, but some inhibitory responses remained even after vagotomy. These results indicate that hypothalamic stimulation induced both excitatory and inhibitory responses in pancreatic exocrine secretion via the vagus and other routes.
Animals↗
In vivo effects by estrone sulfate on the central nervous system-senile dementia (Alzheimer's type).
Seven women with senile dementia-Alzheimer's type (SDAT) were treated with conjugated estrogen [main content: estrone sulfate (E1-S)], at a dose of 1.25 mg/day over a 6-week period. A New Screening Test for Dementia developed by Japanese National Institute of Mental Health (NS) and the scores of Hasegawa Scale for dementia (HS) were performed every 3 weeks. Six women showed improvements in NS (P less than 0.05) and 5 women showed improvements in HS. Untreated women with SDAT did not show any improvement. Serum E1-S was measured by a direct radioimmunoassay. Serum E1-S was 911 +/- 156 pg/ml in 7 women with SDAT and lower than that of 7 normal women (1020 +/- 216 pg/ml). Following the treatment, serum E1-S increased to a level of 21.1 +/- 8.1 ng/ml. Estrone and estradiol-17 beta also increased. The results suggest a possibility for the future clinical use of estrogen for senile dementia, after careful clinical research trials including the side effects.
Aged↗