Heavy chain of human high molecular weight and low molecular weight kininogens binds calcium ion

An antibody subpopulation, anti high molecular weight (anti-HMW) kininogen-Ca2+ antibody able to bind specifically to the HMW kininogen-Ca2+ complex, was isolated from anti-HMW kininogen antiserum. Partially purified anti-HMW kininogen antibody was applied to a HMW kininogen-Sepharose column equilibrated with 40 mM tris(hydroxymethyl)aminomethane hydrochloride buffer, pH 7.5, containing 1.0 M NaCl and 1 mM CaCl2, and anti-HMW kininogen-Ca2+ antibody was eluted with 5 mM ethylenediaminetetraacetic acid. As a result of characterization by enzyme-linked immunosorbent assay, this antibody specifically recognized the cyanogen bromide cleaved fragment 1 (CB-1) region (1-160 amino acid sequence) of the heavy chain of kininogen molecules in the presence of Ca2+ or Mg2+. Furthermore, circular dichroism (CD) experiments showed that the conformational changes of HMW kininogen and heavy chain were induced by metal ions such as Ca2+ and Mg2+ and that these changes were due to the conformational change of the CB-1 region of the heavy chain. The dissociation constant (Kd) for the heavy chain-Ca2+ measured by CD analysis at 214 nm was found to be 0.33 +/- 0.09 mM (mean +/- SD). The number of Ca2+-binding sites of heavy chain calculated from the Hill plot was 1.15 +/- 0.04 (mean +/- SD). Then, a possible Ca2+-binding site was found in the amino-terminal portion of the heavy chain of kininogen molecules.     

Pancreatic B-cell function in non-insulin-dependent diabetes mellitus during successive periods of sulfonylurea and insulin treatment: serum C-peptide response to glucagon and urine C-peptide excretion

Serum C-peptide responses to glucagon and daily urine C-peptide excretion in successive periods of different treatment in two groups of patients with non-insulin-dependent diabetes mellitus (NIDDM) (mean interval between two tests less than 1 month) were compared. In group A patients (n = 8), the glycemic control was improved after transferring the treatment from sulfonylurea (SU) to insulin (fasting plasma glucose: SU: 192 +/- 47, insulin: 127 +/- 21 mg/dl, mean +/- S.D., p less than 0.01). Fasting serum C-peptide immunoreactivity (CPR) was significantly lower at the period of insulin treatment (SU: 1.93 +/- 1.01, insulin: 1.47 +/- 0.79 ng/ml, p less than 0.05), but there was no difference in the increase in serum CPR (maximal--fasting) (delta serum CPR) during glucagon stimulation in the two periods of treatment (SU: 1.70 +/- 0.72, insulin: 1.47 +/- 0.98 ng/ml). In group B patients (n = 7), there was no significant difference in glycemic control after transferring the treatment from insulin to SU (fasting plasma glucose: insulin: 127 +/- 24, SU: 103 +/- 13 mg/dl). Fasting serum CPR was significantly lower during the period of insulin treatment (insulin: 1.39 +/- 0.64, SU: 2.21 +/- 0.86 ng/ml, p less than 0.025), but delta serum CPR during glucagon stimulation still showed no significant difference between the two periods (insulin: 1.97 +/- 1.16, SU: 2.33 +/- 1.57 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)     

Specificity of the interaction of furfural with DNA

Furfural or 2-furaldehyde is a dietary mutagen and is present in various frequently consumed food products. The alkaline unwinding assay and protection of cleavage sites from the action of various restriction enzymes was used to study the interaction of furfural with DNA. Alkaline unwinding experiments showed the formation of an increasing number of strand breaks in duplex DNA both with increasing furfural concentration and with time of reaction. Treatment of lambda phage DNA with furfural protected cleavage with restriction endonucleases DraI and SspI but not with ApaI, BssHII and SacII. These results indicate that under the conditions used furfural reacts exclusively with AT base pairs. A minimum of 3-4 consecutive AT base pairs are required for this reaction. This was determined by the use of several restriction enzymes whose hexanucleotide recognition sequences contain subsets of AT base pairs.     

Interleukin-6 triggers the association of its receptor with a possible signal transducer, gp130

Interleukin-6 mediates pleiotropic functions in various types of cells through its specific receptor (IL-6-R), the cDNA of which has already been cloned. We report here that an 80 kd single polypeptide chain (IL-6-R) is involved in IL-6 binding and that IL-6 triggers the association of this receptor with a non-ligand-binding membrane glycoprotein, gp130. The association takes place at 37 degrees C within 5 min and is stable for at least 40 min in the presence of IL-6, but does not occur at 0 degree C. Human IL-6-R can associate with a murine gp130 homolog and is functional in murine cells. Mutant IL-6-R lacking the intracytoplasmic portion is functional, suggesting that the two polypeptide chains interact to involve their extracellular portion. In fact, a soluble IL-6-R lacking the transmembrane and intracytoplasmic domains can associate with gp130 in the presence of IL-6 and mediate its function. These findings indicate that the complex of IL-6 and IL-6-R can interact with a non-ligand-binding membrane glycoprotein, gp130, extracellularly and can provide the IL-6 signal.     

A game analysis of reciprocal cooperation: Sequential food-sharing and sex role alternation

The evolution of reciprocal cooperation between non-relatives is studied. Food-sharing of vampire bats studied by Wilkinson (1984) and egg-trading of simultaneous hermaphroditic fish studied by Fisher (1980) are respectively considered as Thompson's reciprocal assistance and the iterated Hero game. Those models predicted the following: (1) Reciprocal assistance between adult bats is evolutionarily stable if they repeatedly interact for a long time. However, (2) the cost for an adult to assist a juvenile is too high to be compensated by the benefit from the juvenile. (3) A particular signal, which determines the sex role of each partner, is always displayed by a “heroic” partner which releases eggs with a larger cost. (4) If alternation of sex roles is established, then the evolutionarily stable frequency of displaying a signal of egg-release increases as the frequency of divorce decreases, and (5) a “cheating” strategy, which releases sperm on every spawning bout, is less advantageous than reciprocal cooperation.     

Comparison of antigenic properties of the X-potato virus and its coat proteins. Effect of proteolytic cleavage on antigenic characteristics

Antigenic properties of intact potato virus X (PVX) particles and of PVX coat protein (CP) preparations were compared using different modifications of ELISA test. In the competitive ELISA test (reaction in solution) antibodies to intact virus react much stronger with PVX than with CP while antibodies to CP react much stronger with CP than with PVX. In the direct ELISA test (reaction on the solid support) the difference in reactions of antiCP antibodies with PVX and CP is eliminated while the one in reactions of antiPVX antibodies with these antigens remains. No difference was registered in reactivity of PVX absorbed directly on polystyrene or on immunoglobulin-coated wells (sandwich ELISA) to antiCP antibodies.     

The effect of rimantadine on the structure of model and biological membranes

The action of the antiviral drug rimantadine on the structure of bilayer lipid membranes (BLM) and RBC membranes was investigated. Structural changes in BLM were recorded by ionophore conductivity changes and by changes in the third harmonic of capacity current signal due to lateral compression of BLM in an electric field. It was shown that the adsorption of rimantadine on BLM results in an increase in ionophore mobility in bilayer membranes of dioleolyllecithin (DOL) and common lipids of bovine brain (CL) and in a decrease in those of azolectin (A). Relative changes in the third harmonic signal also depend on the membrane composition and have different signs. The results may be explained by the rimantadine action on the lipid bilayer structure: "rigidification" of A-membranes and "fluidization" of BLM from DOL and CL. Structural reorganization of RBC membranes as investigated by the ability of the cells to enter a micropipette (inner diameter greater than or equal to 3 microns) thereby undergoing deformation. It was shown that rimantadine influences RBC deformability due to drug induced inhomogenous mechanical membrane properties. Also, rimantadine accelerated the process of artificially induced aggregation of erythrocytes. The relation of the effects on artificial and biological membranes, and the structural changes in the lipid phase of membrane are discussed.     

Development of nephrotic ICGN mice--the origin, reproductive ability, and incidence of glomerulonephritis

ICGN is a strain of spontaneous nephrotic mice with nonproliferative glomerular lesions. It was derived from an outbred Yok: ICR colony in our laboratory. The renal disease constantly occurred in animals of the first to the tenth generations (greater than 13.0%; 70 days of age). When affected males were mated with unaffected females, the incidence of the disease in their offspring was 38.8% (n = 49) at 70 days after birth. When both parents were affected, their offspring were all affected (n = 12). The disease evenly progressed in both sexes. It usually began 40 to 150 days after birth and death occurred within two months after onset. The animals usually showed sufficient reproductive ability as long as unaffected females were used for mating.     

The phenobarbital-type induction of rat liver microsomal monooxygenases by perfluorodecalin

Induction of perfluorodecalin (PFD) of the liver microsomal system of metabolism of xenobiotics has been studied and compared with the inductions by phenobarbital (PB) and 3-methylcholanthrene (MC). It has been shown that PFD increases the content of cytochrome P-450, NADPH-cytochrome c reductase activity. Like PB, PFD induces the activities of benzphetamine-N-demethylase, aldrine-epoxidase, 16 beta-androstendion-hydroxylase. Using specific antibodies against cytochromes P-450b and P-450c (which are the main isoenzymes of cytochrome P-450 in the PB- and MC-microsomes respectively), an immunological identity of the cytochrome P-450 isoforms during PFD and PB induction has been found. According to the rocket immunoelectrophoresis the content of cytochrome P-450 in PFD-microsomes, which is immunologically indistinguishable from P-450b, was approximately 70% of the total cytochrome P-450. Two forms of cytochrome P-450 were isolated from the liver microsomes of PFD-induced rats and purified to homogeneity. A comparison of these forms with cytochromes P-450b and P-450e obtained from the PB-induced rat liver microsomes revealed their similarity in a number of properties, e.g., chromotographic behavior on DEAE-Sephacel column, molecular weight determined by sodium dodecyl sulphate (SDS) electrophoresis in polyacrylamide gel, immunoreactivity, peptide mapping, catalytic activity. The data presented demonstrate that PFD induced in rat liver microsomes the cytochrome P-450 forms whose immunological properties and substrate specificity correspond to those of the PB-type cytochrome P-450. These findings suggest that PFD and PB, which differ in their chemical structure, induce in the rat liver microsomes identical forms of cytochrome P-450.     

The ATP/ADP-antiporter is involved in the uncoupling effect of fatty acids on mitochondria

The ATP/ADP-antiporter inhibitors and the substrate ADP suppress the uncoupling effect induced by low (10-20 microM) concentrations of palmitate in mitochondria from skeletal muscle and liver. The inhibitors and ADP are found to (a) inhibit the palmitate-stimulated respiration in the controlled state and (b) increase the membrane potential lowered by palmitate. The degree of efficiency decreases in the order: carboxyatractylate (CAtr) greater than ADP greater than bongkrekic acid, atractylate. GDP is ineffective, Mg.ADP is of much smaller effect, whereas ATP is effective at much higher concentration than is ADP. Inhibitor concentrations, which maximally suppress the palmitate-stimulated respiration, correspond to those needed for arresting the state 3 respiration. The extent of the CAtr-sensitive stimulation of respiration by palmitate has been found to decrease with an increase in palmitate concentration. Stimulation of the controlled respiration by p-trifluoromethoxycarbonylcyanide phenylhydrozone (FCCP) and gramicidin D at any concentrations of these uncouplers is CAtr-insensitive, whereas that caused by a low concentrations of 2,4-dinitrophenol and dodecyl sulfate is inhibited by CAtr. The above effect of palmitate develops immediately after addition of the fatty acid. It is resistant to EGTA as well as to inhibitors of phospholipase (nupercain) and of lipid peroxidation (ionol). Moreover, palmitate accelerates spontaneous release of the respiratory control, developing in rat liver mitochondria under certain conditions. This effect takes several minutes, being sensitive to EGTA, nupercain and ionol. Like the fast uncoupling, this slow effect is inhibited by ADP but CAtr and atractylate are stimulatory rather than inhibitory. In artificial planar phospholipid membrane, palmitate does not increase the membrane conductance, FCCP increases it strongly and dinitrophenol only slightly. In cytochrome oxidase proteoliposomes, FCCP, gramicidin and dinitrophenol (less effectively) lower, whereas palmitate enhances the cytochrome-oxidase-generated membrane potential. In this system, monensin substitutes for palmitate. It is concluded that the ATP/ADP antiporter is somehow involved in the uncoupling effect caused by low concentrations of palmitate and, partially, of dinitrophenol, whereas uncoupling produced by FCCP and gramicidin is due to their action on the phospholipid part of the mitochondrial membrane. A possible mechanism of this effect is discussed.