Botulinum Neurotoxin A4 Has a 1000-Fold Reduced Potency Due to Three Single Amino Acid Alterations in the Protein Receptor Binding Domain
Botulinum neurotoxin subtype A4 (BoNT/A4) is ~1000-fold less potent than BoNT/A1. This study addresses the basis for low BoNT/A4 potency. Utilizing BoNT/A1-A4 and BoNT/A4-A1 Light Chain-Heavy Chain (LC-HC) chimeras, HC-A4 was responsible for low BoNT/A4 potency. Earlier studies showed BoNT/A1-recept...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2023-03-01
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| Series: | International Journal of Molecular Sciences |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1422-0067/24/6/5690 |
| Summary: | Botulinum neurotoxin subtype A4 (BoNT/A4) is ~1000-fold less potent than BoNT/A1. This study addresses the basis for low BoNT/A4 potency. Utilizing BoNT/A1-A4 and BoNT/A4-A1 Light Chain-Heavy Chain (LC-HC) chimeras, HC-A4 was responsible for low BoNT/A4 potency. Earlier studies showed BoNT/A1-receptor binding domain (Hcc) bound a β-strand peptide (556–564) and glycan-N<sup>559</sup> within Luminal Domain 4 (LD4) of SV2C, the BoNT/A protein receptor. Relative to BoNT/A1, the Hcc of BoNT/A4 possesses two amino acid variants (D<sup>1141</sup> and N<sup>1142</sup>) within the β-peptide binding interface and one amino acid variant (R<sup>1292</sup>) located near the SV2C glycan-N<sup>559</sup>. Introduction of BoNT/A4 β-strand peptide variant (D<sup>1141</sup> and N<sup>1142</sup>) into BoNT/A1 reduced toxin potency 30-fold, and additional introduction of the BoNT/A4 glycan-N<sup>559</sup> variant (D<sup>1141</sup>, N<sup>1142</sup>, and R<sup>1292</sup>) further reduced toxin potency to approach BoNT/A4. While introduction of BoNT/A1 glycan-N<sup>559</sup> variant (G<sup>1292</sup>) into BoNT/A4 did not alter toxin potency, additional introduction of BoNT/A1 β-strand peptide variants (G<sup>1141</sup>, S<sup>1142</sup>, and G<sup>1292</sup>) resulted in potency approaching BoNT/A1 potency. Thus, outcomes from these functional and modeling studies indicate that in rodent models, disruption of Hcc -SV2C β-peptide and -glycan-N<sup>559</sup> interactions mediate low BoNT/A4 potency, while in human motor neurons, disruption of Hcc-SV2C β-peptide alone mediates low BoNT/A4 potency, which link to a species-specific variation at SV2C<sup>563</sup>. |
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| ISSN: | 1661-6596 1422-0067 |