Takashi Katsu

Interaction of wasp venom mastoparan with biomembranes

Mastoparan-induced changes in the K+ permeability of rat peritoneal mast cells, human erythrocytes, Staphylococcus aureus and Escherichia coli were examined. Mastoparan did not efficiently increase the K+ permeability of cells except for S. aureus. The release of membrane phospholipids was also observed from S. aureus cells in the concentration range of the permeability enhancement. Mastoparan stimulated histamine release from mast cells, independently of a small efflux of K+. Mastoparan became markedly effective to E. coli cells whose outer membrane structure was chemically disrupted beforehand, showing that the peptide can enhance the permeability of the cytoplasmic membranes of both Gram-positive and -negative bacteria. In experiments using liposomes, mastoparan increased the permeability of the liposomes composed of egg phosphatidylethanolamine and egg phosphatidylglycerol, which are the lipid constituents of the cytoplasmic membrane of E. coli cells, while it showed a weak activity to the liposomes composed of egg phosphatidylcholine and cholesterol. The latter result related closely to the fact that this peptide acted weakly on erythrocytes and mast cells in which acidic lipids constitute a minor portion. Mastoparan decreased the phase transition temperature of dipalmitoylphosphatidylglycerol liposomes, but it did not affect that of dipalmitoylphosphatidylcholine liposomes. These results indicate that mastoparan penetrated into membranes mainly containing acidic phospholipids and disrupted the membrane structure to increase the permeability. The action of the wasp venom mastoparan was compared with that of a bee venom melittin.

Potentiometric method for the determination of adenosine-5′-triphosphate

Abstract A potentiometric method for the determination of adenosine-5′-triphosphate (ATP) concentrations using a choline-sensitive membrane electrode and choline kinase enzyme is described. The choline electrode was constructed from a poly(vinyl chloride)-based membrane containing sodium tetrakis[3,5-bis(2-methoxyhexafluoro-2-propyl)phenyl]borate as an ion exchanger and 2-fluoro-2′-nitrodiphenyl ether as a membrane solvent. Choline reacted quantitatively with ATP to produce the electrode-insensitive choline phosphate and, therefore, the concentration of ATP could be determined by the measurement of choline consumed by the enzymatic reaction. ATP down to 10 μM could be detected. This method was used successfully to determine the intracellular ATP content of human red blood cells.