title: Structural stabilization of botulinum neurotoxins by tyrosine phosphorylation.
abstract: Tyrosine phosphorylation of botulinum neurotoxins augments their proteolytic activity and thermal stability, suggesting a substantial modification of the global protein conformation. We used Fourier-transform infrared (FTIR) spectroscopy to study changes of secondary structure and thermostability of tyrosine phosphorylated botulinum neurotoxins A (BoNT A) and E (BoNT E). Changes in the conformationally-sensitive amide I band upon phosphorylation indicated an increase of the alpha-helical content with a concomitant decrease of less ordered structures such as turns and random coils, and without changes in beta-sheet content. These changes in secondary structure were accompanied by an increase in the residual amide II absorbance band remaining upon H-D exchange, consistent with a tighter packing of the phosphorylated proteins. FTIR and differential scanning calorimetry (DSC) analyses of the denaturation process show that phosphorylated neurotoxins denature at temperatures higher than those required by non-phosphorylated species. These findings indicate that tyrosine phosphorylation induced a transition to higher order and that the more compact structure presumably imparts to the phosphorylated neurotoxins the higher catalytic activity and thermostability.
authors.affiliation: Department of Neurochemistry, University Miguel Hernández, C/Monóvar s/n (Polígono de Carrús), Alicante, Spain.
authors.forename: J A
authors.initials: JA
authors.lastname: Encinar
authors.forename: A
authors.initials: A
authors.lastname: Fernández
authors.forename: J A
authors.initials: JA
authors.lastname: Ferragut
authors.forename: J M
authors.initials: JM
authors.lastname: González-Ros
authors.forename: B R
authors.initials: BR
authors.lastname: DasGupta
authors.forename: M
authors.initials: M
authors.lastname: Montal
authors.forename: A
authors.initials: A
authors.lastname: Ferrer-Montiel
mesh_terms: D001905:Botulinum Toxins / Q000737:chemistry*; D002152:Calorimetry, Differential Scanning; D010766:Phosphorylation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D014443:Tyrosine; D001905:Botulinum Toxins