Immunochemical identification of the ADP-ribosyltransferase in botulinum C1 neurotoxin as C3 exoenzyme - like molecule

Narito Morii, Yasuhiro Ohashi, Yasuo Nemoto, Motohatsu Fujiwara, Yasuchika Ohnishi, Tei-ichi Nishiki, Yoichi Kamata, Shunji Kozaki, Shuh Narumiya, Genji Sakaguchi

Research output: Contribution to journalArticle

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Abstract

Botulinum C1 neurotoxin and C3 exoenzyme were purified to apparent homogeneity from the culture filtrate of Clostridium botulinum type C strain 003-9. Both preparations catalyzed ADP-ribosylation of the same substrate, the Mr 22, 000 rho gene product (Gb). When the light and heavy chains of C1 toxin were separated, ADP-ribosyltransferase activity in the toxin was quantitatively recovered in the light chain fraction. Anti-C1 toxin antiserum precipitated the ADP-ribosyltransferase activity and the neurotoxicity of C1 toxin in parallel, whereas it had no effect on C3 exoenzyme. On the other hand, anti-C3 exoenzyme anti serum precipitated the ADP-ribosyltransferase activities of both C3 exoenzyme and C1 toxin. This antibody, however, did not precipitate the neurotoxicity of C1 toxin. The ADP-ribosyltransferase in C1 toxin was quantitatively adsorbed onto the anti-C3 antibody column and separated from the majority of C1 toxin protein. The enzyme was then eluted with acidic urea and Western blotting analysis of this eluate revealed the appearance of a protein band positively stained with anti-C3 antibody at a position similar to that of C3 exoenzyme. Quantitative determination by enzyme-linked immunosorbent assay showed that the C3-like immunoreactivity is present in the C1 toxin molecules at the molecular ratio of 1 to 1, 000. These results suggest that the ADP-ribosyltransferase activity in Cl toxin is expressed by a C3-like molecule which is present in a small amount in the toxin preparation and appears to bind to the toxin components). The above results also indicate that the ADP-ribosyltransferase in C1 toxin is not related to its neurotoxin action.

Original languageEnglish
Pages (from-to)769-775
Number of pages7
JournalJournal of Biochemistry
Volume107
Issue number5
Publication statusPublished - May 1990
Externally publishedYes

Fingerprint

ADP Ribose Transferases
Administrative data processing
Neurotoxins
Antibody
Molecules
Preparation
Antibodies
Protein
Enzyme-linked Immunosorbent Assay
Anti-Idiotypic Antibodies
Clostridium botulinum type C
Homogeneity
Enzymes
Light
Clostridium
Immunosorbents
Proteins
Substrate
Gene
Adenosine Diphosphate

ASJC Scopus subject areas

  • Statistics, Probability and Uncertainty
  • Applied Mathematics
  • Physiology (medical)
  • Radiology Nuclear Medicine and imaging
  • Molecular Biology
  • Biochemistry

Cite this

Morii, N., Ohashi, Y., Nemoto, Y., Fujiwara, M., Ohnishi, Y., Nishiki, T., ... Sakaguchi, G. (1990). Immunochemical identification of the ADP-ribosyltransferase in botulinum C1 neurotoxin as C3 exoenzyme - like molecule. Journal of Biochemistry, 107(5), 769-775.

Immunochemical identification of the ADP-ribosyltransferase in botulinum C1 neurotoxin as C3 exoenzyme - like molecule. / Morii, Narito; Ohashi, Yasuhiro; Nemoto, Yasuo; Fujiwara, Motohatsu; Ohnishi, Yasuchika; Nishiki, Tei-ichi; Kamata, Yoichi; Kozaki, Shunji; Narumiya, Shuh; Sakaguchi, Genji.

In: Journal of Biochemistry, Vol. 107, No. 5, 05.1990, p. 769-775.

Research output: Contribution to journalArticle

Morii, N, Ohashi, Y, Nemoto, Y, Fujiwara, M, Ohnishi, Y, Nishiki, T, Kamata, Y, Kozaki, S, Narumiya, S & Sakaguchi, G 1990, 'Immunochemical identification of the ADP-ribosyltransferase in botulinum C1 neurotoxin as C3 exoenzyme - like molecule', Journal of Biochemistry, vol. 107, no. 5, pp. 769-775.
Morii, Narito ; Ohashi, Yasuhiro ; Nemoto, Yasuo ; Fujiwara, Motohatsu ; Ohnishi, Yasuchika ; Nishiki, Tei-ichi ; Kamata, Yoichi ; Kozaki, Shunji ; Narumiya, Shuh ; Sakaguchi, Genji. / Immunochemical identification of the ADP-ribosyltransferase in botulinum C1 neurotoxin as C3 exoenzyme - like molecule. In: Journal of Biochemistry. 1990 ; Vol. 107, No. 5. pp. 769-775.
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abstract = "Botulinum C1 neurotoxin and C3 exoenzyme were purified to apparent homogeneity from the culture filtrate of Clostridium botulinum type C strain 003-9. Both preparations catalyzed ADP-ribosylation of the same substrate, the Mr 22, 000 rho gene product (Gb). When the light and heavy chains of C1 toxin were separated, ADP-ribosyltransferase activity in the toxin was quantitatively recovered in the light chain fraction. Anti-C1 toxin antiserum precipitated the ADP-ribosyltransferase activity and the neurotoxicity of C1 toxin in parallel, whereas it had no effect on C3 exoenzyme. On the other hand, anti-C3 exoenzyme anti serum precipitated the ADP-ribosyltransferase activities of both C3 exoenzyme and C1 toxin. This antibody, however, did not precipitate the neurotoxicity of C1 toxin. The ADP-ribosyltransferase in C1 toxin was quantitatively adsorbed onto the anti-C3 antibody column and separated from the majority of C1 toxin protein. The enzyme was then eluted with acidic urea and Western blotting analysis of this eluate revealed the appearance of a protein band positively stained with anti-C3 antibody at a position similar to that of C3 exoenzyme. Quantitative determination by enzyme-linked immunosorbent assay showed that the C3-like immunoreactivity is present in the C1 toxin molecules at the molecular ratio of 1 to 1, 000. These results suggest that the ADP-ribosyltransferase activity in Cl toxin is expressed by a C3-like molecule which is present in a small amount in the toxin preparation and appears to bind to the toxin components). The above results also indicate that the ADP-ribosyltransferase in C1 toxin is not related to its neurotoxin action.",
author = "Narito Morii and Yasuhiro Ohashi and Yasuo Nemoto and Motohatsu Fujiwara and Yasuchika Ohnishi and Tei-ichi Nishiki and Yoichi Kamata and Shunji Kozaki and Shuh Narumiya and Genji Sakaguchi",
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AU - Fujiwara, Motohatsu

AU - Ohnishi, Yasuchika

AU - Nishiki, Tei-ichi

AU - Kamata, Yoichi

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