title: Botulinum neurotoxin G binds synaptotagmin-II in a mode similar to that of serotype B: tyrosine 1186 and lysine 1191 cause its lower affinity.
abstract: Botulinum neurotoxins (BoNTs) block neurotransmitter release by proteolyzing SNARE proteins in peripheral nerve terminals. Entry into neurons occurs subsequent to interaction with gangliosides and a synaptic vesicle protein. Isoforms I and II of synaptotagmin were shown to act as protein receptors for two of the seven BoNT serotypes, BoNT/B and BoNT/G, and for mosaic-type BoNT/DC. BoNT/B and BoNT/G exhibit a homologous binding site for synaptotagmin whose interacting part adopts helical structure upon binding to BoNT/B. Whereas the BoNT/B-synaptotagmin-II interaction has been elucidated in molecular detail, corresponding information about BoNT/G is lacking. Here we systematically mutated the synaptotagmin binding site in BoNT/G and performed a comparative binding analysis with mutants of the cell binding subunit of BoNT/B. The results suggest that synaptotagmin takes the same overall orientation in BoNT/B and BoNT/G governed by the strictly conserved central parts of the toxins' binding site. The surrounding nonconserved areas differently contribute to receptor binding. Reciprocal mutations Y1186W and L1191Y increased the level of binding of BoNT/G approximately to the level of BoNT/B affinity, suggesting a similar synaptotagmin-bound state. The effects of the mutations were confirmed by studying the activity of correspondingly mutated full-length BoNTs. On the basis of these data, molecular modeling experiments were employed to reveal an atomistic model of BoNT/G-synaptotagmin recognition. These data suggest a reduced length and/or a bend in the C-terminal part of the synaptotagmin helix that forms upon contact with BoNT/G as compared with BoNT/B and are in agreement with the data of the mutational analyses.
authors: Institut für Physiologische Chemie, OE 4310.
authors: Gesche
authors: G
authors: Willjes
authors: Stefan
authors: S
authors: Mahrhold
authors: Jasmin
authors: J
authors: Strotmeier
authors: Timo
authors: T
authors: Eichner
authors: Andreas
authors: A
authors: Rummel
authors: Thomas
authors: T
authors: Binz
mesh_terms: D000818:Animals; D001665:Binding Sites; D001905:Botulinum Toxins / Q000235:genetics / Q000378:metabolism* / Q000633:toxicity; D019274:Botulinum Toxins, Type A; D002417:Cattle; D005677:G(M1) Ganglioside / Q000031:analogs & derivatives / Q000378:metabolism; D005732:Gangliosides / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D050863:Synaptotagmin I / Q000235:genetics / Q000378:metabolism; D050861:Synaptotagmin II / Q000235:genetics / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism
chemical_list: D005732:Gangliosides; D050863:Synaptotagmin I; D050861:Synaptotagmin II; C096323:rimabotulinumtoxinB; C064020:ganglioside GD1alpha; C488535:botulinum toxin type G; D005677:G(M1) Ganglioside; D014443:Tyrosine; C016481:trisialoganglioside GT1; D001905:Botulinum Toxins; D019274:Botulinum Toxins, Type A; D008239:Lysine