Crystal structure of the receptor binding domain of the botulinum C–D mosaic neurotoxin reveals potential roles of lysines 1118 and 1136 in membrane interactions

• Published in: Biochemical and biophysical research communications Vol. 404; no. 1; pp. 407 - 412
• Main Authors: Zhang, Yanfeng, Buchko, Garry W., Qin, Ling, Robinson, Howard, Varnum, Susan M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 07.01.2011
• Subjects: AMINO ACID SEQUENCE; amino acid sequences; ANIMALS; bacteria; binding properties; binding sites; Botulinum neurotoxin; botulinum neurotoxin, C-D mosaic, botulism, phosphatidylethanolamine, membrane recognition, SSGCID; Botulinum toxin; Botulinum Toxins - chemistry; Botulinum Toxins - genetics; Botulinum Toxins - metabolism; Botulism; Cell Membrane - metabolism; CHAINS; CLOSTRIDIUM BOTULINUM; Clostridium botulinum C; Clostridium botulinum D; CRYSTAL STRUCTURE; Crystallography, X-Ray; C–D mosaic; DISEASES; Environmental Molecular Sciences Laboratory; Glycolipids; Humans; Hybrids; Infection; LIPOSOMES; Liposomes - chemistry; Lysine; Lysine - chemistry; Lysine - genetics; MATERIALS SCIENCE; Membrane recognition; MEMBRANES; Molecular Sequence Data; Mosaics; Mutation; Neurotoxins; Phosphatidylethanolamine; phosphatidylethanolamines; Phosphatidylethanolamines - chemistry; PHOSPHOLIPIDS; Phospholipids - chemistry; Protein Structure, Tertiary; PROTEINS; RESOLUTION; Salts; Serotypes; STRAINS; Sulfate; SULFATES; Synaptosomes; C–D mosaic; Phosphatidylethanolamine; Hcc; Botulinum neurotoxin; BoNT; Hcn; PC; Botulism; PE; HCR; Lc; BoNT/D-HCR; Hc; Membrane recognition; BoNT/CD-HCR
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2010.11.134

► Crystal structure of botulinum C–D mosaic neurotoxin at 1.56Å resolution. ► BoNT/CD-HCR binds PE liposomes more tightly than BoNT/D-HCR. ► Lysine-1118 may enhance BoNT/CD-HCR synaptosome binding by stabilizing a putative membrane interaction loop. ► Binding of a sulfate ion to K1136 may reflect natural interactions of BoNT/CD-HCR with membrane phospholipids or glycolipids. The botulinum neurotoxins (BoNTs) produced by different strains of the bacterium Clostridium botulinum are responsible for the disease botulism and include a group of immunologically distinct serotypes (A, B, E, and F) that are considered to be the most lethal natural proteins known for humans. Two BoNT serotypes, C and D, while rarely associated with human infection, are responsible for deadly botulism outbreaks afflicting animals. Also associated with animal infections is the BoNT C–D mosaic protein (BoNT/CD), a BoNT subtype that is essentially a hybrid of the BoNT/C (∼two-third) and BoNT/D (∼one-third) serotypes. While the amino acid sequence of the heavy chain receptor binding (HCR) domain of BoNT/CD (BoNT/CD-HCR) is very similar to the corresponding amino acid sequence of BoNT/D, BoNT/CD-HCR binds synaptosome membranes better than BoNT/D-HCR. To obtain structural insights for the different membrane binding properties, the crystal structure of BoNT/CD-HCR (S867-E1280) was determined at 1.56Å resolution and compared to previously reported structures for BoNT/D-HCR. Overall, the BoNT/CD-HCR structure is similar to the two sub-domain organization observed for other BoNT HCRs: an N-terminal jellyroll barrel motif and a C-terminal β-trefoil fold. Comparison of the structure of BoNT/CD-HCR with BoNT/D-HCR indicates that K1118 has a similar structural role as the equivalent residue, E1114, in BoNT/D-HCR, while K1136 has a structurally different role than the equivalent residue, G1132, in BoNT/D-HCR. Lysine-1118 forms a salt bridge with E1247 and may enhance membrane interactions by stabilizing the putative membrane binding loop (K1240-N1248). Lysine-1136 is observed on the surface of the protein. A sulfate ion bound to K1136 may mimic a natural interaction with the negatively changed phospholipid membrane surface. Liposome-binding experiments demonstrate that BoNT/CD-HCR binds phosphatidylethanolamine liposomes more tightly than BoNT/D-HCR.