Proteomic analysis of mammalian basic proteins by liquid-based two-dimensional column chromatography

Yu Kyong Shin, Hyoung Joo Lee, Joon Seok Lee, Young Ki Paik

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

To develop a standard method for separating highly basic proteins in mammalian cells, we established a 2-D LC separation system coupled with chromatofocusing/nonporous RP column chromatography (CF/NPRPC) in a ProteomeLab PF2D system. After standardizing conditions for 2-D LC, a 2-D liquid protein map of uninfected macrophage proteins with pH range 8.3-11.3 was constructed, and then compared with a macrophage protein map made after infection with Candida albicans. The results demonstrate that 2-D LC offers both high resolution and reproducibility for separation of highly basic, macrophage proteins. After protein identification using a nano 2-D LC-MS/MS Proteomics Solution System, quantitative determination of the changes in the differentially expressed proteins (e.g., galectin-3) in C. albicans-infected macrophages was also accomplished by measuring the peak area of the chromatogram in 2-D LC. The result from this measurement of galectin-3 expression shows a 3.41-fold decrease in the infected macrophage cells, which was further confirmed by that from the RT-PCR of mRNA of galectin-3. Thus, 2-D LC coupled with CF/NPRPC could be applicable to common analysis of highly basic proteins in a high-throughput manner.

Original languageEnglish
Pages (from-to)1143-1150
Number of pages8
JournalProteomics
Volume6
Issue number4
DOIs
Publication statusPublished - 2006 Feb 1

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Column chromatography
Proteomics
Chromatography
Macrophages
Liquids
Galectin 3
Proteins
Candida albicans
Candida
Cells
Throughput
Polymerase Chain Reaction
Messenger RNA
Infection

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

Cite this

Shin, Yu Kyong ; Lee, Hyoung Joo ; Lee, Joon Seok ; Paik, Young Ki. / Proteomic analysis of mammalian basic proteins by liquid-based two-dimensional column chromatography. In: Proteomics. 2006 ; Vol. 6, No. 4. pp. 1143-1150.
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Proteomic analysis of mammalian basic proteins by liquid-based two-dimensional column chromatography. / Shin, Yu Kyong; Lee, Hyoung Joo; Lee, Joon Seok; Paik, Young Ki.

In: Proteomics, Vol. 6, No. 4, 01.02.2006, p. 1143-1150.

Research output: Contribution to journalArticle

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