Crystal Structure of k-Bungarotoxin at 2.3-Å Resolution

John C. Dewan, Gregory A. Grant, James Sacchettini

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70 Scopus citations

Abstract

k-Neurotoxins display a very low affinity for neuromuscular receptors, but bind tightly to, and inhibit, nicotinic acetylcholine receptors in neuronal tissue such as the chick ciliary ganglia. In contrast, α-neurotoxins bind with high affinity and inhibit nicotinic acetylcholine receptors at the neuromuscular junction. The origin of this difference in specificity has been a long-studied question in the field. Here we report the first crystal structure of a k-neurotoxin, k-bungarotoxin. Unlike the NMR structure previously reported [Sutcliffe, M. J., Dobson, C. M., & Oswald, R. E. (1992) Biochemistry 31, 2962-2970], the present crystal structure more accurately defines the polypeptide fold and the nature of the interaction between subunits in the active dimer, which is a unique feature of the k-neurotoxins. The structure has been refined to R = 19.6% with X-ray diffraction data extending to a resolution of 2.3 A. There are two independent protein molecules (66 amino acid residues each) in the asymmetric unit that are arranged as a dimer with the two subunits related by a rotation of 178.6°. Each subunit consists of three main-chain loops. Three of the five β-strands of each subunit form an antiparallel β-sheet which becomes an extended six-stranded antiparallel β-sheet, by virtue of the approximate 2-fold symmetry of the dimer. The interactions at the dimer interface consist of six main-chain-main-chain hydrogen bonds, as well as three other hydrogen-bonding interactions involving side chains. Residues Phe 49 and Leu 57 are found in all four k-bungarotoxins that have been sequenced, but occur in no α-neurotoxins, and they form van der Waals interactions across the dimer interface. The two subunits of the dimer are not identical, with the major difference between them occurring at the tip of the central loop (Cys 27-Pro 36). Residue Arg 34, which is essential for the activity of both α- and k-neurotoxins, occurs at the tip of the central loop in each subunit, with guanidinium groups that are ∼44 Å apart.

Original languageEnglish
Pages (from-to)13147-13154
Number of pages8
JournalBiochemistry
Volume33
Issue number44
DOIs
StatePublished - Nov 1 1994

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