Structure of botulinum neurotoxin type D light chain at 1.65 A resolution: repercussions for VAMP-2 substrate specificity.
The seven serotypes (A-G) of botulinum neurotoxins (BoNTs) function through their proteolytic cleavage of one of three proteins (SNAP-25, Syntaxin, and VAMP) that form the SNARE complex required for synaptic vesicle fusion. The different BoNTs have very specific protease recognition requirements, between 15 and 50 amino acids in length depending on the serotype. However, the structural details involved in substrate recognition remain largely unknown. Here is reported the 1.65 A resolution crystal structure of the catalytic domain of BoNT serotype D (BoNT/D-LC), providing insight into the protein-protein binding interaction and final proteolysis of VAMP-2. Structural analysis has identified a hydrophobic pocket potentially involved in substrate recognition of the P1' VAMP residue (Leu 60) and a second remote site for recognition of the V1 SNARE motif that is critical for activity. A structural comparison of BoNT/D-LC with BoNT/F-LC that also recognizes VAMP-2 one residue away from the BoNT/D-LC site provides additional molecular details about the unique serotype specific activities. In particular, BoNT/D prefers a hydrophobic interaction for the V1 motif of VAMP-2, while BoNT/F adopts a more hydrophilic strategy for recognition of the same V1 motif.
Arndt
Joseph W
JW
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Chai
Qing
Q
Christian
Todd
T
Stevens
Raymond C
RC
D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001905:Botulinum Toxins / Q000737:chemistry* / Q000378:metabolism; D003014:Clostridium botulinum / Q000378:metabolism; D018360:Crystallography, X-Ray; D057927:Hydrophobic and Hydrophilic Interactions; D008957:Models, Genetic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D050600:SNARE Proteins / Q000378:metabolism; D012549:Schistosoma japonicum / Q000378:metabolism; D013379:Substrate Specificity; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics / Q000378:metabolism; D050988:Vesicle-Associated Membrane Protein 2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D011993:Recombinant Fusion Proteins; D050600:SNARE Proteins; D050825:Synaptosomal-Associated Protein 25; D050988:Vesicle-Associated Membrane Protein 2; C096325:botulinum toxin type D; D001905:Botulinum Toxins