The isolation and purification of two peptides from the venom of Buthus martensii Karsch

Two peptides that extensively prolong action potentials (APs) in rat and fr og nerves have been isolated and purified from the venom of the scorpion Bu thus martensii Karsch (BMK). The peptides were purified using gel filtratio n, ion exchange, FPLC, and HPLC chromatography. Action potentials recorded in the presence of nanomolar concentrations of the peptides were extensivel y prolonged without much attenuation in their heights. The N-terminal seque nces of both the peptides, BMK 9(3)-1 and BMK 9(3)-2, were determined. The N-terminal sequences of BMK 9(3)-1 and BMK 9(3)-2 were found to be: GRDAYIA DSEN-PYF-GANPN and GRDAYIADSEN-PYT-ALNP. Sequence similarity comparisons to other alpha-scorpion toxins suggest that the two blanks in each of the seq uences are cysteines. The first 20 residues of the two BMK peptides differ by only three amino acid substitutions. The molecular weight (MW) of BMK 9( 3)-1 and BMK 9(3)-2 were determined by LC/MS/MS to be 7020 and 7037 Da. Sin ce both of the peptides prolong APs when both K+ and Ca+ (+) channels are b locked and show sequence similarity to other cc-neurotoxins, it appears lik ely that BMK 9(3)-1 and BMK 9(3)-2 act to alter Na channel inactivation to produce their effects. The first 20 residues of BMK 9(3)-2 are identical to those observed for makaloxin I, a toxin isolated from Buthus martensii Kar sch venom, that alters nitric oxide transmitter release. Since the two toxi ns also have very similar molecular weights, BMK 9(3)-2 may be identical to makatoxin I; however, BMK 9(3)-2 acts to alter Na channels to exert its ef fect, thus the two toxins may differ, or if they are identical, they can ex ert effects on both neural transmission and AP propagation. (C) 2000 Elsevi er Science Ltd. All rights reserved.