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Structure Analysis of Activity-dependent Neurotrophic Factor 9 by Circular Dichroism and Sedimentation Equilibrium

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Abstract

Activity-dependent neurotrophic factor (ADNF) is a glia-derived neurotrophic peptide, which protects neurons from tetrodoxin treatment and Alzheimer’s disease-related and amyotrophic-lateral-sclerosis-related insults at femto-molar concentrations. However, the mechanism of the femto-molar neuroprotection by the peptide has not been elucidated. The characterization of the peptide structure in solution at molecular level should shed light in the mechanism of such extremely high biological activity. From that point of view, the secondary and quaternary structure analysis of ADNF9, an active core fragment peptide of ADNF, was performed by circular dichroism (CD) and sedimentation equilibrium. ADNF9 has also been shown to exhibit a neurotrophic activity in femto-molar concentrations; in this study it showed sub-pM neuroprotective activity against V642I-APP-induced cytotoxity in the mouse primary cortical neuron. CD analysis showed that the secondary structure of ADNF9 is identical in water and phosphate-buffered saline (PBS) and is independent of the peptide concentration. The CD spectra appear to be characterized most likely as disordered. The sedimentation equilibrium experiments demonstrated monomeric structure of the protein over the wide range of peptide concentration. There is a slight enhancement of CD intensity at 37°C relative to 20°C, suggesting a possible hydrophobic association of the peptide. There is no change in the secondary structure in PBS upon freeze–thaw treatment, which has previously been suggested to cause activity loss.

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Acknowledgments

This study was supported by grants from Japan Society for the Promotion of Science Grant-in-Aid for Science Research (C) 17590893 (Y.K.), 17590894 (T.N.).

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Author notes

  1. Takako Niikura

    Present address: Department of Neurology, Georgetown University, Washington, DC, 20057, USA

  2. Hirohisa Tajima

    Present address: Department of Pharmacology, Tokyo Medical University, 6-1-1 Shinjyuku, Shinjyuku-ku, Tokyo, 160-8402, Japan

Authors and Affiliations

  1. Alliance Protein Laboratories, 3957 Corte Cancion, Thousand Oaks, California, 91360, USA

    Tsutomu Arakawa

  2. Department of Pharmacology, KEIO University School of Medicine, 35 Shinanomachi, Shinjyuku-ku, Tokyo, 160-8582, Japan

    Takako Niikura, Hirohisa Tajima & Yoshiko Kita

  3. Department of Molecular Bioprocessing, Tokyo Institute of Technology, Yokohama, Japan

    Fumio Arisaka

Authors
  1. Tsutomu Arakawa
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  2. Takako Niikura
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  3. Hirohisa Tajima
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  4. Fumio Arisaka
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  5. Yoshiko Kita
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Corresponding author

Correspondence to Yoshiko Kita.

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Arakawa, T., Niikura, T., Tajima, H. et al. Structure Analysis of Activity-dependent Neurotrophic Factor 9 by Circular Dichroism and Sedimentation Equilibrium. J Mol Neurosci 33, 262–267 (2007). https://doi.org/10.1007/s12031-007-0032-5

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  • DOI: https://doi.org/10.1007/s12031-007-0032-5

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