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Title: Structural and mutational analyses of the receptor binding domain of botulinum D/C mosaic neurotoxin: Insight into the ganglioside binding mechanism

Abstract

Highlights: {yields} We determined the crystal structure of the receptor binding domain of BoNT in complex with 3'-sialyllactose. {yields} An electron density derived from the 3'-sialyllactose was confirmed at the cleft in the C-terminal subdomain. {yields} Alanine site-directed mutagenesis showed that GBS and GBL are important for ganglioside binding. {yields} A cell binding mechanism, which involves cooperative contribution of two sites, was proposed. -- Abstract: Clostridium botulinum type D strain OFD05, which produces the D/C mosaic neurotoxin, was isolated from cattle killed by the recent botulism outbreak in Japan. The D/C mosaic neurotoxin is the most toxic of the botulinum neurotoxins (BoNT) characterized to date. Here, we determined the crystal structure of the receptor binding domain of BoNT from strain OFD05 in complex with 3'-sialyllactose at a resolution of 3.0 A. In the structure, an electron density derived from the 3'-sialyllactose was confirmed at the cleft in the C-terminal subdomain. Alanine site-directed mutagenesis showed the significant contribution of the residues surrounding the cleft to ganglioside recognition. In addition, a loop adjoining the cleft also plays an important role in ganglioside recognition. In contrast, little effect was observed when the residues located around the surface previously identified as the protein receptormore » binding site in other BoNTs were substituted. The results of cell binding analysis of the mutants were significantly correlated with the ganglioside binding properties. Based on these observations, a cell binding mechanism of BoNT from strain OFD05 is proposed, which involves cooperative contribution of two ganglioside binding sites.« less

Authors:
 [1];  [2];  [3]; ;  [4]; ;  [5];  [1];  [3];  [1];  [3]
  1. Graduate School of Life Sciences, Hokkaido University, Sapporo 060-0810 (Japan)
  2. Creative Research Institution 'Sousei,' Hokkaido University, Sapporo 001-0021 (Japan)
  3. (Japan)
  4. Department of Microbiology, Fujita Health University School of Medicine, Toyoake, Aichi 470-1192 (Japan)
  5. Department of Veterinary Science, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka 598-8531 (Japan)
Publication Date:
OSTI Identifier:
22207421
Resource Type:
Journal Article
Journal Name:
Biochemical and Biophysical Research Communications
Additional Journal Information:
Journal Volume: 411; Journal Issue: 2; Other Information: Copyright (c) 2011 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0006-291X
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ALANINES; CATTLE; CLOSTRIDIUM BOTULINUM; CRYSTAL STRUCTURE; ELECTRON DENSITY; MUTAGENESIS; RECEPTORS; TOXICITY

Citation Formats

Nuemket, Nipawan, Tanaka, Yoshikazu, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Tsukamoto, Kentaro, Tsuji, Takao, Nakamura, Keiji, Kozaki, Shunji, Yao, Min, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Tanaka, Isao, E-mail: tanaka@castor.sci.hokudai.ac.jp, and Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810. Structural and mutational analyses of the receptor binding domain of botulinum D/C mosaic neurotoxin: Insight into the ganglioside binding mechanism. United States: N. p., 2011. Web. doi:10.1016/J.BBRC.2011.06.173.
Nuemket, Nipawan, Tanaka, Yoshikazu, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Tsukamoto, Kentaro, Tsuji, Takao, Nakamura, Keiji, Kozaki, Shunji, Yao, Min, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Tanaka, Isao, E-mail: tanaka@castor.sci.hokudai.ac.jp, & Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810. Structural and mutational analyses of the receptor binding domain of botulinum D/C mosaic neurotoxin: Insight into the ganglioside binding mechanism. United States. doi:10.1016/J.BBRC.2011.06.173.
Nuemket, Nipawan, Tanaka, Yoshikazu, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Tsukamoto, Kentaro, Tsuji, Takao, Nakamura, Keiji, Kozaki, Shunji, Yao, Min, Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810, Tanaka, Isao, E-mail: tanaka@castor.sci.hokudai.ac.jp, and Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810. Fri . "Structural and mutational analyses of the receptor binding domain of botulinum D/C mosaic neurotoxin: Insight into the ganglioside binding mechanism". United States. doi:10.1016/J.BBRC.2011.06.173.
@article{osti_22207421,
title = {Structural and mutational analyses of the receptor binding domain of botulinum D/C mosaic neurotoxin: Insight into the ganglioside binding mechanism},
author = {Nuemket, Nipawan and Tanaka, Yoshikazu and Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810 and Tsukamoto, Kentaro and Tsuji, Takao and Nakamura, Keiji and Kozaki, Shunji and Yao, Min and Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810 and Tanaka, Isao, E-mail: tanaka@castor.sci.hokudai.ac.jp and Faculty of Advanced Life Science, Hokkaido University, Sapporo 060-0810},
abstractNote = {Highlights: {yields} We determined the crystal structure of the receptor binding domain of BoNT in complex with 3'-sialyllactose. {yields} An electron density derived from the 3'-sialyllactose was confirmed at the cleft in the C-terminal subdomain. {yields} Alanine site-directed mutagenesis showed that GBS and GBL are important for ganglioside binding. {yields} A cell binding mechanism, which involves cooperative contribution of two sites, was proposed. -- Abstract: Clostridium botulinum type D strain OFD05, which produces the D/C mosaic neurotoxin, was isolated from cattle killed by the recent botulism outbreak in Japan. The D/C mosaic neurotoxin is the most toxic of the botulinum neurotoxins (BoNT) characterized to date. Here, we determined the crystal structure of the receptor binding domain of BoNT from strain OFD05 in complex with 3'-sialyllactose at a resolution of 3.0 A. In the structure, an electron density derived from the 3'-sialyllactose was confirmed at the cleft in the C-terminal subdomain. Alanine site-directed mutagenesis showed the significant contribution of the residues surrounding the cleft to ganglioside recognition. In addition, a loop adjoining the cleft also plays an important role in ganglioside recognition. In contrast, little effect was observed when the residues located around the surface previously identified as the protein receptor binding site in other BoNTs were substituted. The results of cell binding analysis of the mutants were significantly correlated with the ganglioside binding properties. Based on these observations, a cell binding mechanism of BoNT from strain OFD05 is proposed, which involves cooperative contribution of two ganglioside binding sites.},
doi = {10.1016/J.BBRC.2011.06.173},
journal = {Biochemical and Biophysical Research Communications},
issn = {0006-291X},
number = 2,
volume = 411,
place = {United States},
year = {2011},
month = {7}
}