Structure of apamin in solution: a two-dimensional nuclear magnetic resonance study.

A two-dimensional (2-D) Fourier-transform nuclear magnetic resonance (NMR) study of the 18 amino acid neurotoxin apamin isolated from honeybee venom is reported. Combining 2-D J-correlated spectra with 2-D nuclear Overhauser spectra in H2O solution permits essentially complete assignment of the 1H NMR spectrum of apamin at a fixed pH, including a number of spin systems that are reported for the first time. The structural model previously derived by Bystrov et al. [Bystrov, V. F., Okhanov, V. V., Miroshnikov, A. I., & Ovchinnikov, Yu. A. (1980) FEBS Lett. 119, 113-116] from NMR data is shown to be largely correct. The 2-D nuclear Overhauser effect (NOE) spectrum in particular reveals a series of amide-amide NOE's consistent with an alpha-helical core (residues 9-15) in the molecule. NOE's between amide and C alpha protons, followed by amide to amide NOE's, indicate a beta-turn involving residues 3-5 and a nonstandard turn including residues 6-8. We find no evidence for the beta-type structure postulated at the C terminus, however. Instead, it appears that the alpha-helix continues with increasing fraying from residues 16-18.