Huwentoxin-Ⅺ purified from the Chinese bird spider Oithoctonus huwena is a toxin with both antiprotease activity and potassium channel blocking activity. To determine its solution structure,huwentoxin-Ⅺ was expressed in a yeast eukaryotic expression system and studied by NMR. The 15N labeling strategy was used to facilitate the process of resonance assignments. The nearly complete sequence-specific assignments of proton and nitrogen resonances were obtained by analyzing a series of two-dimensional (2D)and three-dimensional (3D) spectra, including DQF-COSY, TOCSY, NOESY, 15N-1H HSQC, 15N-1H HNHA,15N-1H HNHB, 15N-1H TOCSY-HSQC and 15N-1H NOESY-HSQC spectra. Secondary structure analysis of huwentoxin-Ⅺ showed that it mainly contains an N-terminal 310-helix from Thr3 to Arg5 and a C-terminal α-helix from Gln45 to Cys52, plus a triple-stranded antiparallel β-sheet of Glu18-Asn23, Thr26-Ile31 and Asn40-Lys41. These studies provide a solid basis for the final structure determination of huwentoxin-Ⅺ.