This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Clostridial neurotoxins (CNTs), such as botulinum (BoNT) and tetanus (TeNT) neurotoxins, are the causative agents of the neuroparalytic diseases tetanus and botulism. CNTs impair neuronal exocytosis by specific proteolysis of SNARE proteins once inside the neuron, resulting in the clinical manifestations of flaccid and spastic motor paralysis. CNTs bind with high specificity at neuromuscular junctions. The molecular details of the toxin-cell recognition have been elusive. We reported the structure of a BoNT in complex with its protein receptor: the receptor-binding domain of botulinum neurotoxin serotype B (BoNT/B) bound to the luminal domain of synaptotagmin II, determined at 2.15-? resolution. On binding, a helix is induced in the luminal domain that binds to a saddle-shaped crevice on a distal tip of BoNT/B. This crevice is adjacent to the non-overlapping ganglioside-binding site of BoNT/B. Biochemical and neuronal ex vivo studies of structure-based mutations indicate high specificity and affinity of the interaction, and high selectivity of BoNT/B among synaptotagmin I and II isoforms. Synergistic binding of both synaptotagmin and ganglioside imposes geometric restrictions on the initiation of BoNT/B translocation after endocytosis. The mechanism by which a CNT properly identifies and cleaves its target SNARE once inside the neuron involves one or more regions of enzyme-substrate interaction remote from the active site (exosites). Our studies provide the basis for the development of preventive vaccines or inhibitors against these neurotoxins for bio defense, as well as design of modified neurotoxins with different target specificities for clinical applications. In addition, this work is a paradigm for protease inhibitor development in general since proteases represent major challenges for drug development due to the inherent flexibility of these enzymes.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001209-32
Application #
8362050
Study Section
Special Emphasis Panel (ZRG1-BCMB-P (40))
Project Start
2011-03-01
Project End
2012-02-29
Budget Start
2011-03-01
Budget End
2012-02-29
Support Year
32
Fiscal Year
2011
Total Cost
$8,489
Indirect Cost
Name
Stanford University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
Remesh, Soumya G; Andreatta, Massimo; Ying, Ge et al. (2017) Unconventional Peptide Presentation by Major Histocompatibility Complex (MHC) Class I Allele HLA-A*02:01: BREAKING CONFINEMENT. J Biol Chem 292:5262-5270
VanderLinden, Ryan T; Hemmis, Casey W; Yao, Tingting et al. (2017) Structure and energetics of pairwise interactions between proteasome subunits RPN2, RPN13, and ubiquitin clarify a substrate recruitment mechanism. J Biol Chem 292:9493-9504
Zhang, Haonan; Qiao, Anna; Yang, Dehua et al. (2017) Structure of the full-length glucagon class B G-protein-coupled receptor. Nature 546:259-264
Niedzialkowska, Ewa; Mruga?a, Beata; Rugor, Agnieszka et al. (2017) Optimization of overexpression of a chaperone protein of steroid C25 dehydrogenase for biochemical and biophysical characterization. Protein Expr Purif 134:47-62
de Vries, Robert P; Tzarum, Netanel; Peng, Wenjie et al. (2017) A single mutation in Taiwanese H6N1 influenza hemagglutinin switches binding to human-type receptors. EMBO Mol Med 9:1314-1325
Tolbert, William D; Gohain, Neelakshi; Alsahafi, Nirmin et al. (2017) Targeting the Late Stage of HIV-1 Entry for Antibody-Dependent Cellular Cytotoxicity: Structural Basis for Env Epitopes in the C11 Region. Structure 25:1719-1731.e4
Yoon, Chun Hong; DeMirci, Hasan; Sierra, Raymond G et al. (2017) Se-SAD serial femtosecond crystallography datasets from selenobiotinyl-streptavidin. Sci Data 4:170055
Warelow, Thomas P; Pushie, M Jake; Cotelesage, Julien J H et al. (2017) The active site structure and catalytic mechanism of arsenite oxidase. Sci Rep 7:1757
Tzarum, Netanel; de Vries, Robert P; Peng, Wenjie et al. (2017) The 150-Loop Restricts the Host Specificity of Human H10N8 Influenza Virus. Cell Rep 19:235-245
Hettle, Andrew; Fillo, Alexander; Abe, Kento et al. (2017) Properties of a family 56 carbohydrate-binding module and its role in the recognition and hydrolysis of ?-1,3-glucan. J Biol Chem 292:16955-16968

Showing the most recent 10 out of 581 publications