CHANGING THE STRUCTURAL CONTEXT OF A FUNCTIONAL BETA-HAIRPIN - SYNTHESIS AND CHARACTERIZATION OF A CHIMERA CONTAINING THE CURAREMIMETIC LOOP OF A SNAKE TOXIN IN THE SCORPION ALPHA BETA SCAFFOLD/
E. Drakopoulou et al., CHANGING THE STRUCTURAL CONTEXT OF A FUNCTIONAL BETA-HAIRPIN - SYNTHESIS AND CHARACTERIZATION OF A CHIMERA CONTAINING THE CURAREMIMETIC LOOP OF A SNAKE TOXIN IN THE SCORPION ALPHA BETA SCAFFOLD/, The Journal of biological chemistry, 271(20), 1996, pp. 11979-11987
An approach to obtain new active proteins is the incorporation of all
or a part of a well defined active site onto a natural structure actin
g as a structural scaffold. According to this strategy we tentatively
engineered a new curaremimetic molecule by transferring the functional
central loop of a snake toxin, sequence 26-37, sandwiched between two
hairpins, onto the structurally similar beta-hairpin of the scorpion
toxin charybdotoxin, stabilized by a short helix. The resulting chimer
ic molecule, only 31 amino acids long, was produced by solid phase syn
thesis, refolded, and purified to homogeneity. As shown by structural
analysis performed by CD and NMR spectroscopy, the chimera maintained
the expected alpha/beta fold characteristic of scorpion toxins and pre
sented a remarkable structural stability. The chimera competitively di
splaces the snake curaremimetic toxin alpha from the acetylcholine rec
eptor at 10(-5) M concentrations. Antibodies, elicited in rabbits agai
nst the chimera, recognize the parent snake toxin and prevent its bind
ing to the acetylcholine receptor, thus neutralizing its toxic functio
n. All these data demonstrate that the strategy of active site transfe
r to the charybdotoxin scaffold has general applications in the engine
ering of novel ligands for membrane receptors and in vaccine design.