Absence Of Receptor Protein Research Articles

Clostridium perfringens type A enterotoxin forms mepacrine-sensitive pores in pure phospholipid bilayers in the absence of putative receptor proteins

Clostridium perfringens enterotoxin (CPE) is an important cause of food poisoning with no significant homology to other enterotoxins and its mechanism of action remains uncertain. Although CPE has recently been shown to complex with tight junction proteins, we have previously demonstrated that CPE increases ionic permeability in single Caco-2 cells using the whole-cell patch-clamp technique, thereby excluding any paracellular permeability. In this paper we demonstrate that CPE forms pores in synthetic phospholipid membranes in the absence of receptor proteins. The properties of the pores are consistent with CPE-induced permeability changes in Caco-2 cells suggesting that CPE has innate pore-forming ability.

Membrane damage of liposomes by the mushroom toxin phallolysin

We have investigated the membrane-damaging effect of phallolysin on liposomes varying in phospholipid composition, net charge and physical constitution. Liposomes were prepared from lipids extracted from bovine or human erythrocyte ghosts. The liposomes composed of bovine lipids (the intact cell showing little sensitivity to phallolysin) were found comparably sensitive to those prepared from lipids of human red cells (these cells being of high sensitivity). In addition, artificial mixtures of lipids were used for the preparation of liposomes, consisting of (a) negatively charged phospholipids such as dicetyl phosphate or phosphatidylserine, (b) cholesterol, and (c) either sphingomyelin (as the major component of erythrocytes from ruminants) or phosphatidylcholine (as the major component of erythrocytes from non-ruminants). Again, we found only little difference in the susceptibilities of sphingomyelin- and phosphatidylcholine-containing liposomes. On the other hand, the susceptibility depended on the presence of phospholipids with negative net charges. Omittance of phosphatidylcholine or dicetyl phosphate, or replacement by the positively charged stearylamine, decreased the susceptibility by a factor of more than 20. Finally, we prepared liposomes from dicetyl phosphate, cholesterol and phosphatidylcholine in two physical states: large unilamellar and smaller multilamellar liposomes. The unilamellar liposomes were about 10-times more sensitive to phallolysin. We conclude: (1) Phallolysin damages phospholipid-membranes in the absence of receptor proteins, but high concentrations of the toxin are required. (2) Membrane damage takes place with liposomes containing phosphatidylcholine as well as those containing sphingomyelin. (3) Phallolysin damages only liposomes containing phospholipids with a negative net charge.

Isoelectric focusing of oestradiol receptor protein from human mammary carcinoma—A comparison with dextran coated charcoal assay

Two simple methods for the detection of specific oestradiol—17β receptor protein in breast tumour tissue were investigated. Isoelectric focusing of receptor protein in polyacrylamide gel was examined as an alternative micromethod to dextran-coated charcoal receptor assay. In a series of 32 cytosols prepared from unselected breast tumour tissue, assay results agreed on the presence or absence of receptors in 97% of cases. Quantitative analysis of the positive results showed a correlation coefficient, r = 0.88. Isoelectric focusing gave significantly lower receptor concentrations than the dextran-coated charcoal method. Twenty-six Tru-Cut biopsy tumour specimens were analyzed for receptor by isoelectric focusing and the results were compared with those obtained by the dextran-coated charcoal method on the same tumour. In 88% of the specimens examined, both methods agreed as to the presence or absence of receptor protein. A correlation coefficient, r = 0.62. was calculated from the 15 positive results. There was no significant difference between the results from the two methods. Quantitative results were related to total protein concentration in the cytosol. Negative receptor status was defined as less than 5 fmol/mg cytosol protein for both methods.