title: Evidence against formation of A23187 dimers and oligomers in solution: photo-induced degradation of Ionophore A23187.
abstract: Ionophore A23187 has been proposed to form Ca(2+)- conducting channels that arise from dimers and oligomers of the compound (e.g., Balasubramanian, S. V., and Easwaran, K. R. K. (1989) Biochem. Biophys. Res. Commun. 158, 891-897). To investigate this possibility, the solution behavior of A23187 in chloroform, n-hexane, ethanol, 80% methanol-water, and palmitoyloleoylphosphatidyl choline (POPC) vesicles was investigated using UV-VIS, circular dichroism (CD), and 1H NMR techniques. The concentration dependence of the UV-VIS and CD spectra obtained in freshly prepared chloroform solutions indicates that neutral A23187 (HA) exists as a monomer for ionophore concentrations in the range of 50-1000 microM. The cause of time- and concentration-dependent spectral alterations which gave rise to the dimer/channel hypothesis was also investigated. For solutions of 50-1000 microM A23187 in chloroform, n-hexane, and ethanol stored in the dark, no spectral changes were observed for periods of 2 months. However, solutions in these solvents did show time-dependent spectral changes when exposed to light. In 80% methanol-water or phospholipid vesicles, similar spectral changes were observed, even when the solutions were protected from light. Application of TLC and MS methods indicate that the time-dependent spectral changes reflect degradation of A23187, not dimer or oligomer formation. The degradative processes proceed with half-lives ranging from approximately 75 to > 400 h, and are influenced by several factors, including solvent, exposure to light, ionophore concentration, pH, and the presence of metal ions, EDTA, dissolved oxygen, and a radical inhibitor. The kinetics of Ca2+ transport into Quin-2-loaded POPC vesicles by authentic A23187 give no evidence of a channel mechanism, even following a previous and lengthy coincubation of the ionophore with the vesicles.
authors.affiliation: Department of Chemistry and Biochemistry, University of Oklahoma, Norman 73019, USA.
authors.forename: T P
authors.initials: TP
authors.lastname: Thomas
authors.forename: E
authors.initials: E
authors.lastname: Wang
authors.forename: D R
authors.initials: DR
authors.lastname: Pfeiffer
authors.forename: R W
authors.initials: RW
authors.lastname: Taylor
mesh_terms: D000001:Calcimycin / Q000737:chemistry* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002725:Chloroform; D002942:Circular Dichroism; D019281:Dimerization; D006860:Hydrogen Bonding; D007473:Ion Channels / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008027:Light; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008968:Molecular Conformation; D015394:Molecular Structure; D010782:Photolysis; D012997:Solvents; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet
chemical_list: D007473:Ion Channels; D012997:Solvents; D000001:Calcimycin; D002725:Chloroform; D002118:Calcium