Localization of the phalloidin and nucleotide-binding sites on actin.
Phalloidin was found to block nucleotide exchange in F-actin, without interfering with nucleotide hydrolysis. This inhibition of nucleotide exchange occurs under conditions in which monomers are able to exchange. The distance separating a fluorescent chromophore attached to phalloidin from the nucleotide on actin was determined using fluorescence resonance energy-transfer spectroscopy. They are separated by less than 1.0 nm. Added confirmation of the close proximity of phalloidin to nucleotide was obtained by extracting a small peptide-ATP complex from an actin digest. The peptide comprises residues 114-118, which are from the same region as the residues that others have shown to crosslink to phalloidin [Vandekerckhove et al. (1985) EMBO J. 4, 2815-2818]. The results suggest that phalloidin has two major effects. It traps actin monomers in a conformation which appears to be distinct from G-actin and it stabilizes the structure of F-actin, an event accompanied by the trapping of ADP.
D000199:Actins / Q000032:analysis*; D000244:Adenosine Diphosphate / Q000032:analysis; D000255:Adenosine Triphosphate / Q000032:analysis; D000494:Allosteric Regulation; D001665:Binding Sites; D004735:Energy Transfer; D009682:Magnetic Resonance Spectroscopy; D009711:Nucleotides / Q000032:analysis*; D009842:Oligopeptides / Q000032:analysis*; D010446:Peptide Fragments / Q000032:analysis; D010590:Phalloidine / Q000032:analysis*; D013050:Spectrometry, Fluorescence
D000199:Actins; D009711:Nucleotides; D009842:Oligopeptides; D010446:Peptide Fragments; D010590:Phalloidine; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate