THE CRYSTAL STRUCTURE OF CARDIOTOXIN V FROM TAIWAN COBRA VENOM AT 2.19 Å RESOLUTION: ROLE OF WATER BINDING LOOP IN THE FORMATION OF MEMBRANE-BINDING SITE OF P-TYPE CARDIOTOXINS. Chwan-Deng Hsiao¹, Yuh-Ju Sun¹, Wen-guey Wu², Chien-Min Chiang², A-Yen Hsin¹,Crystallography Laboratory, Institute of Molecular Biology Academia Sinica, Taipei, Taiwan 11529^1, Structural Biology Group, Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan 30043²

The crystal structure of cardiotoxin V from Taiwan cobra venom (CTX A5) has been solved at pH 8.5. The refined model shows dimeric assembly and the global monomeric structure is found to be similar to that determined by NMR at pH 3.7. Nevertheless, local conformational differences are detected at two functionally important regions of loop I and II. The first difference between the NMR and X-ray structure of CTX A5 is detected near the tip of loop I and can be attributed to the different protonation state of His-4 at different pH. The second difference, detected at the tip of loop II, is due to the interaction of water with amino acid residues in the loop II region of the cardiotoxin containing Pro-31 (P-type CTX). This interaction forces the normally tapering flexible loop II into a more rigid [[Omega]] shape by bridging the main chain at 27 and 34 positions. Thus a continuous hydrophobic column capable of penetrating the membrane lipid bilayers is formed by the tips of three-finger toxin. These results provide a structural basis for the pH-dependent lipid binding activity of CTXs. In addition, a new membrane-spanning element other than helical and [[beta]]-barrel structure is proposed by the hydrophobic loops of [[beta]]-sheet polypeptides. Also discussed is a model of CTX cation channels to explain the cell lysis and depolarization activity.