Automated phosphoproteome analysis for cultured cancer cells by two-dimensional nanoLC-MS using a calcined titania/C18 biphasic column

Koshi Imami, Naoyuki Sugiyama, Yutaka Kyono, Masaru Tomita, Yasushi Ishihama

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Abstract

We have developed an on-line automated system for phosphoproteome analysis using titania-based phosphopeptide enrichment followed by nanoLC-MS/MS. Titania beads were prepared by calcination of commercial chromatographic titania beads at 800°C to convert the crystalline structure. The obtained rutile-form titania exhibited higher selectivity in phosphopeptide enrichment than commercial titania, even in the absence of a competitive chelating reagent for non-phosphopeptides. For phosphoproteome analysis of human cervical cancer HeLa cells, tryptic digests of the cell extracts were directly injected into this on-line system, and 696 non-redundant phosphopeptides with 671 unambiguously determined phosphorylation sites, derived from 512 phosphoproteins, were successfully identified. This is the first successful application of an on-line automated phosphoproteome analysis system to complex biological samples. 2008

Original languageEnglish
Pages (from-to)161-166
Number of pages6
JournalAnalytical Sciences
Volume24
Issue number1
DOIs
Publication statusPublished - 2008 Jan

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Cells
Phosphopeptides
Phosphorylation
Online systems
Phosphoproteins
Chelation
Calcination
titanium dioxide
Crystalline materials

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Automated phosphoproteome analysis for cultured cancer cells by two-dimensional nanoLC-MS using a calcined titania/C18 biphasic column. / Imami, Koshi; Sugiyama, Naoyuki; Kyono, Yutaka; Tomita, Masaru; Ishihama, Yasushi.

In: Analytical Sciences, Vol. 24, No. 1, 01.2008, p. 161-166.

Research output: Contribution to journalArticle

Imami, Koshi ; Sugiyama, Naoyuki ; Kyono, Yutaka ; Tomita, Masaru ; Ishihama, Yasushi. / Automated phosphoproteome analysis for cultured cancer cells by two-dimensional nanoLC-MS using a calcined titania/C18 biphasic column. In: Analytical Sciences. 2008 ; Vol. 24, No. 1. pp. 161-166.
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