Self-association of band-protein from human erythrocyte membranes in aqueous solutions

Band 3, the main integral protein of the human erythrocyte membrane, was solubilized and purified in high concentrations of acetic acid. After removal of the organic solvent by dialysis, the self-association of the protein in aqueous solutions was studied by analytical ultracentrifugation. Sedimentation velocity and sedimentation equilibrium experiments clearly demonstrate that, under appropriate conditions of protein preparation, at protein concentrations c less than 200 micrograms/ml, ionic strengths 2 less than 10mM and pH values remote from the isoelectric pH of the protein, band 3 shows a monomer/dimer/tetramer-association equilibrium. With some preparations, as well as at higher values of c or I, hexamers and octamers contribute to the association equilibrium. The time needed for relaxation towards association equilibrium depends on the blood donor from whom the membranes were derived and varies between less than one minute and more than several hours. The results of analytical ultracentriguation, together with previously published data on the incorporation of band 3 into planar lipid bilayers, from chemical crosslinking and from electronmicroscopy suggest that band 3 will also show a monomer/dimer/tetramer-association equilibrium in the human erythrocyte membrane. This hypothesis contrasts the widely-held assumption that, in the membrane, band 3 is a stable dimer; however, it is consistent with nearly all known data on band 3-self-association.     

Effects of lanthanum on calcium-dependent phenomena in human red cells.

Lanthanum (0.25 mM) does not penetrate into fresh or Mg2+-depleted cells, whereas it does into ATP-depleted or ATP + 2,3-diphosphoglycerate-depleted cells, into cells containing more than 3 mM calcium, or cells stored for more than 4 weeks in acid/citrate/dextrose solution. In fresh cells loaded with calcium, extracellular lanthanum blocks the active Ca2+-efflux completely and inhibits (Ca2+ + Mg2+)-ATPase (ATP phosphohydrolase, EC 3.6.1.3) activity to about 50%. In Mg2+-depleted cells Ca2+-Ca2+ exchange is inhibited by lanthanum. Ca2+-leak is unaffected by lanthanum up to 0.25 mM concentration; higher lanthanum concentrations reduce leak rate. In NaCl medium Ca2+-leak +/ S.D. amounts to 0.28 +/ 0.08 mumol/1 of cells per min, whereas in KC1 medium to 0.15 +/ 0.04 mumol/1 of cells per min at 2.5 mM [Ca2+]e and 0.25 mM [La3+]e pH 7.1. Lanthanum inhibits Ca2+-dependent rapid K+ transport in ATP-depleted and propranolol-treated red cells, i.e. whenever intracellular calcium is below a critical level. The inhibition of the rapid K+ transport can be attributed to protein-lanthanum interactions on the cell surface, since lanthanum is effectively detached from the membrane lipids by propranolol. Lanthanum at 0.2--0.25 mM concentration has no direct effect on the morphology of red cells. The shape regeneration of Ca2+-loaded cells, however, is blocked by lanthanum owing to Ca2+-pump inhibition. Using lanthanum the transition in cell shape can be quantitatively correlated to intracellular Ca2+ concentrations.     

Influence of the hormone analogue 13-NLE-molitin and of 1-methyl-3-isobutylxanthine on tone and cyclic 3',5'-AMP content of antral and duodenal muscles in the rabbit.

1. By the action of 1-methyl-3-isobutylxanthine (isobutyltheophylline, 2 - 3 × 10⁻⁴ M), the content of cyclic 3', 5'-AMP in the antral and duodenal muscles of the rabbit is increased by 72 % and 126 %, respectively; by 1.8 × 10⁻⁷ M 13-norleucine-motilin and 1.8 × 10⁻⁶ M acetylcholine it is not changed. 13-norleucine-motilin is an analogue of the recently discovered duodenal tissue hormone motilin and has identical effects. 1-methyl-3-isobutylxanthine has a more powerful inhibiting effect on phosphodiesterase than has theophylline.2. 3 × 10⁻⁴ M isobutyltheophylline reduces the tone of the duodenal muscle while simultaneously increasing the content of cyclic AMP and negates the tone-enhancing effect of nle-motilin on the duodenal muscle, while nle-motilin increases the muscle tone lowered by isobutyltheophylline.3. The basic tone of the antral muscle is not reduced by isobutyltheophylline. However, the contraction-promoting effect of nle-motilin after an increase in cyclic AMP due to isobutyltheophylline is significantly lower.4. It is assumed that the changes in the tone or in the response of the antral and duodenal muscles to nle-motilin observed after the administration of isobutyltheophylline, are due to the increase of cyclic AMP in the tissue.5. The antagonistic effects of cyclic AMP and motilin on the gastro-intestinal muscles might be of physiological importance for the regulation of the gastro-intestinal motor activity.     

Non-random intrachromosomal distribution of chromatid aberrations induced by X-rays,alkylating agents and ethanol in Vicia faba

A reconstructed karyotype of Vicia faba with all chromosomes individually distinguishable was treated with triethylene melamine (TEM), cytostasan (CYT) (a new benzimidazol nitrogen mustard), mitomycin C (MI), ethanol (EA) and X-rays. The distribution within chromosomes of induced chromatid abberations was non-random for all agents. The number of segments involved in aberration clustering corresponded to the number of sites representing constitutive heterochromatin, or the regions immediately adjacent to these, as evidenced by the position of Giemsa marker bands. Which of these potential regions of aberration clustering reacted with preferential involvement in aberrations was, in part at least, dependent upon the inducing agent used. It is argued that this may be due to differences in the base composition and/or molecular conformation of heterochromatic regions. Unexpectedly, the distribution pattern of chromatid aberrations induced by mitomycin C was found to be different from those after treatment with the alkylating agents TEM and cytostasan although mitomycin C is assumed to induce aberrations via alkylation. If mitomycin C-induced aberrations are indeed due to alkylation, this indicates that different alkylating agents do not necessarily result in identical patterns of abberation clustering. The other two alkylating agents and ethanol resulted in similar patterns of preferential distribution of abberations. X-Ray induced chromatid aberrations also showed a non-random intrachromosomal distribution, but the clustering was less pronounced than after treatment with the chemical agents.     

Observations on the fine structure,enzyme histochemistry,and innervation of parathyroid gland and ultimobranchial body of Chthonerpeton indistinctum (Gymnophiona,Amphibia)

Summary Fine structural and enzyme histochemical observations on ultimobranchial body and parathyroid gland of the caecilian Chthonerpeton are presented. The cell clusters and follicles of the ultimobranchial body consist mainly of granulated cells which are termed C-cells and obviously belong to the APUD cell series. In the larger follicles additional possibly exhausted degranulated cells and replacement cells occur. A rich supply of nerve fibres has been found in this gland. Frequently nerve terminals were observed to come into synaptic contact with the C-cells. Two categories of nerve fibres occur: a) fibres containing large polymorphic electron dense granules (probably purinergic fibres), b) fibres containing small electron transparent vesicles and a few electron dense granules (probably cholinergic fibres). The parathyroid gland consists of elongated cells (one cell type) poor in organelles and often containing fields of glycogen and lipid droplets. The cells are further characterized by fair amounts of lysosomal enzymes; they are interconnected by maculae adhaerentes and occludentes. No nerves and blood vessels have been found in the parathyroid gland of Chthonerpeton. This study has been supported by the Deutsche Forschungsgemeinschaft We 380/5.     

Estrogen biosynthesis and its inhibition--a review

A short review is given about extragonadal sources of estrogens via androgen aromatization; the possible importance of this aromatization; the mechanism of aromatization, the inhibition of androgen aromatization by steroidal compounds; the importance and possible usefulness of such an inhibition.