Three-Dimensional Structure of Toxin OSK1 from Orthochirus scrobiculosus Scorpion Venom^,

Abstract

A 600 MHz 1H NMR study of toxin OSK1, blocker of small-conductance Ca2+-activated K+ channels, is presented. The unambiguous sequential assignment of all the protons of the toxin was obtained using TOCSY, DQF-COSY, and NOESY experiments at pH 3.0 (10, 30, and 45 degrees C) in aqueous solution. 3J(N alpha), 3J(alphabeta) vicinal spin coupling constants were determined in high-resolution spectra. The cross-peak volumes in NOESY spectra and the coupling constants were used to define the local structure of the protein by the program HABAS and to generate torsion angle and interproton distance constraints for the program DIANA. Hydrogen-deuterium exchange rates of amide protons showed possible locations of hydrogen bonds. The hydrogen bond acceptors and disulfide bridges between residues 8-28, 14-33, and 18-35 were determined when analyzing distance distribution in preliminary DIANA structures. All constraints were used to obtain a set of 30 structures by DIANA. The resulting rms deviations over 30 structures are 1.30 A for the heavy atoms and 0.42 A for the backbone heavy atoms. The structures were refined by constrained energy minimization using the SYBYL program. Their analysis indicated the existence of the alpha-helix (residues 10-21) slightly distorted at the Cys14 residue, two main strands of the antiparallel beta-sheet (24-29, 32-38), and the extended fragment (2-6). The motif is stabilized by the disulfide bridges in the way, common to all known scorpion toxins. Using the fine spatial toxin structure, alignment of the homologues, mutagenesis analysis, and comparison of scorpion toxin family functions, we delineate some differences significant for the toxin specificity.

Pharmacological Profiling of Orthochirus scrobiculosus Toxin 1 Analogs with a Trimmed N-Terminal Domain

“…OSK1 is a toxin that originates from the venom of the Asian scorpion Orthochirus scrobiculosus (Jaravine et al, 1997). It is composed of 38 amino acid residues with three disulfide bridges that are arranged according to a conventional pattern of the type C1-C4, C2-C5, and C3-C6 (Jaravine et al, 1997;Mouhat et al, 2004a,b).…”

“…It is composed of 38 amino acid residues with three disulfide bridges that are arranged according to a conventional pattern of the type C1-C4, C2-C5, and C3-C6 (Jaravine et al, 1997;Mouhat et al, 2004a,b). OSK1, also referred to as ␣-KTx3.7, belongs to the ␣-KTx3 family (Tytgat et al, 1999;Rodriguez de la Vega and Possani, 2004) and shares between 74 and 90% sequence identity with other members of this family, such as kaliotoxin 1 (Aiyar et al, 1995), agitoxins 1, 2, and 3 (Garcia et al, 1994), and Buthus martensi kaliotoxin-like BmkTX (Romi-Lebrun et al, 1997).…”

“…OSK1, also referred to as ␣-KTx3.7, belongs to the ␣-KTx3 family (Tytgat et al, 1999;Rodriguez de la Vega and Possani, 2004) and shares between 74 and 90% sequence identity with other members of this family, such as kaliotoxin 1 (Aiyar et al, 1995), agitoxins 1, 2, and 3 (Garcia et al, 1994), and Buthus martensi kaliotoxin-like BmkTX (Romi-Lebrun et al, 1997). The 3-D solution structure of OSK1, as determined by 1 H NMR (Protein Data Bank accession code 1SCO), indicates that the toxin folds according to the canonical ␣/␤ architectural motif, which corresponds to a helical structure connected to an antiparallel ␤-sheet by two disulfide bridges (Bontems et al, 1991;Jaravine et al, 1997;Mouhat et al, 2004b). In the case of OSK1, the helical structure is a distorted ␣-helix running from amino acid residues 10 to 21, whereas the antiparallel ␤-sheet is composed of two strands going from residues 24 to 28 and 32 to 38.…”

Scorpion sting envenomation presenting with pulmonary edema & subconjunctival hemorrhage: A case report

“…This venom is a species specific complex mixture of short chain neurotoxic proteins, serotonin, hyaluronidase and various enzymes that act on trypsinogen 8,9 . The toxin bind at cell membrane level to the voltage gated K + channels, Ca 2+ activated K + channels and Na + channels.…”

“…The toxin bind at cell membrane level to the voltage gated K + channels, Ca 2+ activated K + channels and Na + channels. 9 IJBAR (2014) 05 (08) www.ssjournals.com…”