Botulinum neurotoxin A changes conformation upon binding to ganglioside GT1b.
In this work, the kinetics of the binding of botulinum neurotoxin A (BoNT/A) to ganglioside GT1b were studied using surface plasmon resonance (SPR). The neurotoxin bound polysialylated gangliosides, and that binding was affected by the ionic strength of the buffer. Although the level of binding was decreased at higher ionic strengths, it could be easily observed in Tris buffer, containing 150 mM NaCl. Data analysis revealed that the binding of BoNT/A to a GT1b-containing phospholipid monolayer did not fit a traditional 1:1 model. Subsequent studies, in which the time of contact between BoNT/A and GT1b was varied, indicated that the BoNT/A-GT1b complex became more stable over time, as evidenced by its reduced rate of dissociation. Circular dichroism indicated that when BoNT/A was incubated with GT1b, it underwent a conformational change that resulted in an increase in alpha-helix content and a decrease in beta-sheet content. Therefore, the SPR kinetic data were fit to a conformational change model and kinetic rate constants determined. The apparent K(D) values obtained for the binding of BoNT/A to ganglioside GT1b ranged from 2.83 x 10(-7) to 1.86 x 10(-7) M, depending on the ionic strength of the buffer.
Department of Biochemistry and Molecular Biology, H171, Pennsylvania State College of Medicine, Hershey, Pennsylvania 17033, USA.
Brian C
BC
Yowler
Cara-Lynne
CL
Schengrund
D019274:Botulinum Toxins, Type A / Q000737:chemistry* / Q000378:metabolism*; D002021:Buffers; D002942:Circular Dichroism; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry; D005732:Gangliosides / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008081:Liposomes; D009994:Osmolar Concentration; D011485:Protein Binding; D011487:Protein Conformation; D012965:Sodium Chloride / Q000737:chemistry; D020349:Surface Plasmon Resonance; D014325:Tromethamine / Q000737:chemistry
D002021:Buffers; D005732:Gangliosides; D008081:Liposomes; D014325:Tromethamine; D012965:Sodium Chloride; C016481:trisialoganglioside GT1; D019274:Botulinum Toxins, Type A; D004134:Dimyristoylphosphatidylcholine