Abstract
Botulinum neurotoxins (BoNT) cause the potentially fatal neuroparalytic disease botulism that arises due to proteolysis of a SNARE protein. Each BoNT is comprised of three domains: a cell binding domain (HC), a translocation domain (HN), and a catalytic (Zn2+ endopeptidase) domain (LC). The HC is responsible for neuronal specificity by targeting both a protein and ganglioside receptor at the neuromuscular junction. Although highly toxic, some BoNTs are commercially available as therapeutics for the treatment of a range of neuromuscular conditions. Here we present the crystal structures of two BoNT cell binding domains, HC/A4 and HC/A5, in a complex with the oligosaccharide of ganglioside, GD1a and GM1b, respectively. These structures, along with a detailed comparison with the previously reported apo-structures, reveal the conformational changes that occur upon ganglioside binding and the interactions involved.
Highlights
Botulinum neurotoxin serotype A (BoNT/A) is produced by anaerobic spore forming bacteria, Clostridium botulinum, and, along with other serotypes, is responsible for the disease botulism—a neuromuscular condition that causes flaccid paralysis and can lead to death by asphyxiation if left untreated [1]
A total of nine hydrogen bonding interactions were present between heavy chain (HC)/A4 and GD1a (Figure 1B) (Table 2)—there was clear electron density for the two terminal nitrogen atoms of Arg 1282 which interact with Sia5 and Gln 1276
The crystal structures of HC/A4:GD1a and HC/A5:GM1b presented here reveal the interactions involved with ganglioside binding and the conformational changes that occur
Summary
Botulinum neurotoxin serotype A (BoNT/A) is produced by anaerobic spore forming bacteria, Clostridium botulinum, and, along with other serotypes, is responsible for the disease botulism—a neuromuscular condition that causes flaccid paralysis and can lead to death by asphyxiation if left untreated [1]. The mechanism of intoxication involves three general steps [5]: highly specific targeting to the neuromuscular junction by dual-receptor recognition of both a protein and ganglioside receptor by the HC domain, resulting in endocytic internalisation into an endosome [6]; pH-mediated conformational change of the HN domain that translocates the LC into the cytosol [7,8,9]; and a Zn2+-dependent endopeptidase cleavage of a soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein by the LC [10] This cleavage prevents vesicular fusion to the cell membrane, halting the release of presynaptic acetylcholine, and the progression of synaptic signalling at the neuromuscular junction [11]
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