Skip to main content
SpringerLink
Log in

Phosphopeptide-selective Column-switching RP-HPLC with a Titania Precolumn

  • Published:
Analytical Sciences Aims and scope Submit manuscript

Abstract

A methodology of phosphopeptide-selective analysis coupled with column-switching HPLC utilizing titania as precolumn media is presented. Phosphopeptides were selectively enriched on titania packing within a protein/peptide mixture without any additional procedure, and analyzed by column-switching high-performance liquid chromatography. First, phospho-compounds were separated from complex mixtures by trapping them under acidic conditions on a titania packing, where non-phosphorylated compounds were effused out of the precolumn. Subsequently, phospho-compounds were desorbed from the titania column under a specific condition and analyzed. The behavior of phospho-compounds on a titania surface, especially adsorption/desorption, was precisely examined and optimized. A phosphoric buffer was successively employed for the elution of phosphopeptides on a titania surface by competition with the free phosphate group. From the successes of a selective concentration/analysis of phosphopeptides with column-switching HPLC with a titania precolumn, a novel phosphopeptide-selective RP-HPLC analysis has been shown to have an application possibility as a tool for phosphoproteomics.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. N. Kawada, D. B. Kristensen, K. Asahina, K. Nakatani, Y. Minamiyama, S. Seki, and K. Yoshizato, J. Biol. Chem., 2001, 276, 25318.

    Article  CAS  Google Scholar 

  2. D. B. Kristensen, N. Kawada, K. Imamura, Y. Miyamoto, C. Tateno, S. Seki, T. Kuroki, and K. Yoshizato, J. Hepatol., 2000, 32, 268.

    Article  CAS  Google Scholar 

  3. Y. Oda, T. Nagasu, and B. T. Chait, Nat. Biotech., 2001, 19, 379.

    Article  CAS  Google Scholar 

  4. H. Zhou, J. D. Watts, and R. Aebersold, Nat. Biotech., 2001, 19, 375.

    Article  CAS  Google Scholar 

  5. C. E. Haydon, P. A. Eyers, L. D. Aveline-Wolf, K. A. Resing, J. L. Maller, and N. G. Ahn, Mol. Cell. Proteomics, 2003, 27.

    Google Scholar 

  6. A. C. Posewitz and P. Tempst, Anal. Chem., 1999, 71, 2883.

    Article  CAS  Google Scholar 

  7. A. Stensballe, S. Andersen, and O. N. Jensen, Proteomics, 2001, 1, 207.

    Article  CAS  Google Scholar 

  8. A. Sickmann and H. E. Meyer, Proteomics, 2001, 1, 200.

    Article  CAS  Google Scholar 

  9. D. A. Sanders, B. L. Gillece-Castro, A. M. Stock, A. L. Burlingame, and D. E. Koshland, Jr., J. Biol. Chem., 1989, 264, 21770.

    Article  CAS  Google Scholar 

  10. T. M. Martensen, Methods Enzymol., 1971, 107, 1.

    Google Scholar 

  11. K. Tani and M. Ozawa, J. Liq. Chromatogr. Rel. Technol., 1999, 22, 843.

    Article  CAS  Google Scholar 

  12. H. Matsuda, H. Nakamura, and T. Nakajima, Anal. Sci., 1990, 6, 911.

    Article  CAS  Google Scholar 

  13. Y. Ikeguchi and H. Nakamura, Anal. Sci., 1997, 13, 479.

    Article  CAS  Google Scholar 

  14. S. Deshimaru, Y. Miyake, T. Ohmiya, Y. Tatsu, Y. Endo, N. Yumoto, and T. Toraya, J. Biochem., 2002, 131, 705.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. GL Sciences Inc., 237–2 Sayamagahara, Iruma, Saitama, 358-0032, Japan

    Ikuma Kuroda, Yukihiro Shintani, Masanori Motokawa, Shuzo Abe & Masahiro Furuno

Authors
  1. Ikuma Kuroda
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yukihiro Shintani
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Masanori Motokawa
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Shuzo Abe
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Masahiro Furuno
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ikuma Kuroda.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kuroda, I., Shintani, Y., Motokawa, M. et al. Phosphopeptide-selective Column-switching RP-HPLC with a Titania Precolumn. ANAL. SCI. 20, 1313–1319 (2004). https://doi.org/10.2116/analsci.20.1313

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.2116/analsci.20.1313

Search

Navigation