Paul K. Pallaghy, Stephen A. Monks and Raymond S. Norton
NMR Laboratory, Biomolecular Research Institute, 381 Royal Parade, Parkville 3052, Australia.
Anthopleurins-A & -B (AP-A & -B), from the sea anemone Anthopleura xanthogrammica, are members of a class of sea anemone toxins which interact specifically with the voltage-gated sodium channel of excitable tissue. In addition to their use as tools in the study of these channels, AP-A and AP-B are potential lead compounds in the development of novel cardiac stimulants due to their potent positive inotropic effect (increased force of cardiac contraction). We have determined their three-dimensional structures in solution by NMR techniques.
AP-A & AP-B, which differ by substitutions at 7 amino acid residues, have essentially identical tertiary folds, comprising a four-stranded anti-parallel -sheet and several -turns. Spectral analysis proved difficult due to the existence of multiple conformations arising from cis-trans isomerisation about the Gly40-Pro41 peptide bond. Differences between AP-A and AP-B occur at the start of a poorly-defined loop connecting the first two -strands. The extent to which differences between the structures correlated with differences in the NMR data, such as chemical shifts, NOE intensities and amide exchange rates, will be discussed.
Some of the residues important for activity have been identified from previous chemical modification and site-directed mutagenesis studies, and the structures allow a corresponding cardioactive pharmacophore to be proposed. The pharmacophore proposed consists of several charged residues including two aspartate residues.
1. Pallaghy, P.K., Scanlon M.J., Monks, S.A. and Norton, R.S. (1995) Biochemistry 34, 3782-3794.
2. Monks, S.A., Pallaghy, P.K., Scanlon, M.J. and Norton, R.S., submitted.