The atomic resolution structure of bucandin, a novel toxin isolated from the Malayan krait, determined by direct methods

Abstract

Bucandin is a novel presynaptic neurotoxin isolated from Bungarus candidus (Malayan krait). It has the unique property of enhancing presynaptic acetylcholine release and represents a family of three-finger toxins with an additional disulfide in the first loop. There are no existing structures from this sub-category of three-finger toxins. The X-ray crystal structure of bucandin has been determined by the Shake-and-Bake direct-methods procedure. The resulting electron-density maps were of outstanding quality and allowed the automated tracing of 61 of the 63 amino-acid residues, including their side chains, and the placement of 48 solvent molecules. The 0.97 A resolution full-matrix least-squares refinement converged to a crystallographic R factor of 12.4% and the final model contains 118 solvent molecules. This is the highest resolution structure of any member of the three-finger toxin family and thus it can serve as the best model for other members of the family. Furthermore, the structure of this novel toxin will help in understanding its unique ability to enhance acetylcholine release. The unique structure resulting from the fifth disulfide bond residing in the first loop improves the understanding of other toxins with a similar arrangement of disulfide bonds.

NMR structure of bucandin, a neurotoxin from the venom of the Malayan krait (Bungarus candidus)

“…The disulphide linkages were identified from medium-resolution structures obtained after three rounds of NOAH calculations. These were found to be consistent with those identified earlier [10] and were used as additional constraints in the subsequent structure calculations. In the final structure calculations 6000 DYANA structures were generated from random starting conformations.…”

“…In the NMR structure, this short strand is situated on the side of the two faces and is slightly offset from the plane defined by the three strands of the second β-sheet. In the X-ray structure, the β-strand is longer, spanning residues 45-48 and is oriented basically on the same plane as the second β-sheet [10].…”

“…The recent atomic-resolution X-ray structure of bucandin shows a highly polar molecule that incorporates two anti-parallel β-sheets [10]. The first loop is isolated from the rest of the molecule, with the tip of the loop twisted away from the rest of the molecule.…”

Solvent reaction field potential inside an uncharged globular protein: A bridge between implicit and explicit solvent models?

“…Structure 1f94 [65], the bucadin snake venom toxin, was selected from the Protein Data Bank [66] as model system for analyzing protein hydration computationally due to its high solubility, compact shape, and highly charged surface residues. To neutralize the overall charge, one of the surface residues, lysine 14, was mutated to phenylalanine.…”

“…To examine the behavior of polar solvent around an uncharged biomolecule, hundreds of nanosecond-scale molecular dynamics simulations were carried out on a fixed conformation of the bucadin snake venom toxin (PDB ID 1f94) [65] immersed in 5926 SPC/E mobile water molecules [19]. Partial charges on all protein atoms were set to zero.…”