Carboxymethylation of the human estrogen receptor ligand-binding domain-estradiol complex: HPLC/ESMS peptide mapping shows that cysteine 447 does not react with iodoacetic acid.

Experiments were carried out to determine the degree of solvent and reagent accessibility of the cysteines in the ligand-binding domain of the human estrogen receptor (hER LBD). The cysteine residues were alkylated when human ER LBD was present in its ligand (estradiol)-bound conformation. Direct electrospray ionization mass spectrometry (ESMS) as well as liquid chromatography coupled with ESMS, and matrix-assisted laser ionization desorption time-of-flight mass spectrometry were used to determine the location and the yield of the derivatized residues after proteolysis with trypsin. We observed that the cysteine 447 was protected against alkylation under these conditions, whereas cysteines 381, 417, and 530 were fully derivatized.

The nature of receptor binding of oxytocin in myometrial tissue.

The dissociation course of the hormone receptor complexes of rat myometrial oxytocin receptors was studied. The dissociation course was biphasic with a major component displaying an almost irreversible binding character (time constant 0.003 min-1). This irreversibility was independent of hormonal status of the myometrium as well as the type of divalent cation present in the medium, and was not caused by endocytosis of the receptor. However, as addition of EDTA caused an immediate dissociation of all receptor binding, the irreversibility must be of a non-covalent character. It is suggested that new analysis methods for receptor kinetics including the possibility of partly irreversible binding mechanisms should be considered.

Regulation of the immune response in Plasmodium falciparum malaria. I. Non-specific proliferative responses in vitro and characterization of lymphocytes.

The mitogen-induced DNA synthesis in vitro in lymphocytes from 20 patients acutely ill with Plasmodium falciparum malaria was compared with that of 16 healthy donors. Within both groups part of the donors were individuals who had only experienced short exposure or none at all to the parasite (Sweden) while the other part were donors living in a malaria endemic area (Colombia). The proliferative response to the T cell mitogen La (leucoagglutinin from PHA) of the patients was significantly reduced as compared with that of the controls. With pokeweed mitogen which stimulates T cells and induces a T cell-dependent activation of B cells, no difference between patients or controls was seen. The results were similar for the donors of different geographical origin and malaria background. Lymphocytes and monocytes from the peripheral blood of these donors were also studied for surface marker distribution by means of monoclonal antibodies. Both the absolute and the relative frequencies of T cells in the blood of the malaria patients were significantly reduced as compared with the controls. Furthermore, in almost all eight patients tested, the ratio between T4+ T cells (including the helper/inducer subsets) and T8+ T cells (including the suppressor and cytotoxic subsets) were below 1:1 while they were close to 2:1 in the controls. The results indicate that the relative frequency of T8+ T cells, expressed as percentage total T cells (T3+) was significantly elevated in the P. falciparum patients. The possible relationship between this imbalance and the irregular La response of the patients lymphocytes requires further investigation of lymphocyte function.