Botulinum Neurotoxin F Subtypes Cleaving the VAMP-2 Q58–K59 Peptide Bond Exhibit Unique Catalytic Properties and Substrate Specificities

Toxins

Mirjam Weil¹  BrigitteG. Dorner¹  MartinB. Dorner¹  Martin Skiba¹  Bazbek Davletov²  Thomas Binz³  Stefan Sikorra³  Jasmin Weisemann⁴  Andreas Rummel⁴  Sylvia Valdezate⁵ 

[1] Biological Toxins (ZBS 3), Centre for Biological Threats and Special Pathogens, Robert Koch Institute, 13353 Berlin, Germany;Department of Biomedical Science, University of Sheffield, Western Bank, Sheffield S10 2TN, UK;Institute of Cell Biochemistry, OE 4310, Hannover Medical School, Carl-Neuberg-Straße 1, 30625 Hannover, Germany;Institute of Toxicology, OE 5340, Hannover Medical School, Carl-Neuberg-Straße 1, 30625 Hannover, Germany;Reference and Research Laboratory for Taxonomy, Spanish National Centre of Microbiology, Institute of Health Carlos III, 28220 Madrid, Spain;

关键词: Clostridium botulinum;    botulinum neurotoxin;    serotype F;    subtype;    vesicle associated membrane protein 2 (VAMP-2);    synaptobrevin;    Zn2+ protease;   

DOI  :  10.3390/toxins10080311

来源: DOAJ

【 摘 要 】

In the recent past, about 40 botulinum neurotoxin (BoNT) subtypes belonging to serotypes A, B, E, and F pathogenic to humans were identified among hundreds of independent isolates. BoNTs are the etiological factors of botulism and represent potential bioweapons; however, they are also recognized pharmaceuticals for the efficient counteraction of hyperactive nerve terminals in a variety of human diseases. The detailed biochemical characterization of subtypes as the basis for development of suitable countermeasures and possible novel therapeutic applications is lagging behind the increase in new subtypes. Here, we report the primary structure of a ninth subtype of BoNT/F. Its amino-acid sequence diverges by at least 8.4% at the holotoxin and 13.4% at the enzymatic domain level from all other known BoNT/F subtypes. We found that BoNT/F9 shares the scissile Q58/K59 bond in its substrate vesicle associated membrane protein 2 with the prototype BoNT/F1. Comparative biochemical analyses of four BoNT/F enzymatic domains showed that the catalytic efficiencies decrease in the order F1 > F7 > F9 > F6, and vary by up to a factor of eight. KM values increase in the order F1 > F9 > F6 ≈ F7, whereas kcat decreases in the order F7 > F1 > F9 > F6. Comparative substrate scanning mutagenesis studies revealed a unique pattern of crucial substrate residues for each subtype. Based upon structural coordinates of F1 bound to an inhibitor polypeptide, the mutational analyses suggest different substrate interactions in the substrate binding channel of each subtype.

【 授权许可】

Unknown