1. Sapecin B, a novel fly toxin, blocks macroscopic K+ currents in the GH3 rat pituitary cell line
N Suzuki, M Hirono, K Kawahara, T Yoshioka Am J Physiol. 1997 Jul;273(1 Pt 1):C289-96. doi: 10.1152/ajpcell.1997.273.1.C289.
Sapecin B is structurally homologous to charybdotoxin (CTX), which is found in scorpion venom. This study investigated the effects of sapecin B on the Ca(2+)-activated K+ currents [IK(Ca)] and the rapidly inactivating K+ currents in clonal rat GH3 pituitary cells with whole cell voltage-clamp methods. Sapecin B (20 nM) reversibly blocked the CTX-sensitive Ix(Ca) (the BK current) in a dose-dependent manner, with a half-maximal inhibitory concentration of approximately 0.9 nM, comparable to that of 0.08-0.4 nM for CTX. The Ca2+ currents in GH3 cells, however, were not affected by sapecin B (40 nM), indicating that the blockade of IK(Ca) by sapecin B is not a secondary effect of Ca2+ current inhibition. The effect of sapecin B on IK(Ca) resembled that of CTX, as expected from the structural similarities shared by CTX and sapecin B. We also found that sapecin B largely inhibited the 4-aminopyridine-sensitive, rapidly inactivating K+ currents in a dose-dependent manner, with a half-maximal inhibitory concentration of approximately 40 nM, whereas CTX had little effect on this current in GH3 cells. Sapecin B may thus provide a useful tool, complementary to CTX, for probing the functional role of molecular domains in the BK channels and the structural similarities common to the BK and the rapidly inactivating A-type K+ channels.
2. Synthesis and characterization of sapecin and sapecin B
J I Kim, H Iwai, S Kurata, M Takahashi, K Masuda, I Shimada, S Natori, Y Arata, K Sato FEBS Lett. 1994 Apr 4;342(2):189-92. doi: 10.1016/0014-5793(94)80498-2.
Two insect defencins, sapecin and sapecin B, were chemically synthesized to confirm their structure and antibacterial activity and also to examine the possibility that these peptides bind to the same site on the large conductance calcium-activated potassium channel as charybdotoxin. Both synthetic peptides showed the same antibacterial activity as native sapecins, indicating that the synthetic peptides folded correctly in the chemical synthesis. Synthetic sapecins did not show an inhibitory effect on [125I]charybdotoxin binding to rat brain synaptic membranes, suggesting that sapecin B recognizes a different binding site from that of charybdotoxin despite the similar structural motif.
3. Purification, sequence and antibacterial activity of two novel sapecin homologues from Sarcophaga embryonic cells: similarity of sapecin B to charybdotoxin
K Yamada, S Natori Biochem J. 1993 Apr 1;291 ( Pt 1)(Pt 1):275-9. doi: 10.1042/bj2910275.
Two sapecin homologues were purified from the culture medium of NIH-Sape-4, an embryonic cell line of Sarcophaga peregrina (flesh fly). These homologues contained six cysteine residues with exactly the same disulphide pairings as those in sapecin. The amino acid sequence of one of them, sapecin C, was also very similar to that of sapecin. The other homologue, sapecin B, was less similar to sapecin but showed significant similarity to charybdotoxin, an inhibitor of K+ channels isolated from a scorpion venom. Like sapecin, these homologues repressed the growth of various Gram-positive bacteria.
