Tài liệu Báo cáo khoa học: Applications of diagonal chromatography for proteomewide characterization

MINIREVIEW

Applications of diagonal chromatography for proteome￾wide characterization of protein modifications and

activity-based analyses

Kris Gevaert1,2, Francis Impens1,2, Petra Van Damme1,2, Bart Ghesquie`re1,2, Xavier Hanoulle3

and Joe¨l Vandekerckhove1,2

1 Department of Medical Protein Research, VIB, Ghent, Belgium

2 Department of Biochemistry, Ghent University, Belgium

3 UMR 8576 CNRS ) University of Sciences and Technologies of Lille, Structural and Functional Glycobiology Unit, Villeneuve d’Ascq,

France

Introduction

Proteomics refers to a qualitative, differential and

quantitative estimation of a proteome. Proteomes can

be extremely complex, often encompassing more than

10 000 different components per cell. Two-dimensional

gel electrophoresis [1] followed by electroblotting and

microsequencing [2–4] or in-gel digestion combined

with Edman sequencing [5] of the generated peptides

or peptide mass fingerprinting [6–10] have been the

methods of choice to reproducibly separate and iden￾tify complex protein mixtures. Although large-scale 2D

gel electrophoresis separates thousands of proteins

[11,12], probably no more than a few hundred different

proteins have been identified from such gels. To obtain

better proteome coverage, alternative methods were

introduced. Groundbreaking methodologies became

available when high-throughput genome sequencing

started to cover the entire genetic information of

several species. This information is now available for a

Keywords

activity-based probe; ATP-binding proteins;

COFRADIC; diagonal chromatography;

N-terminal peptides; peptide sorting; protein

N-glycosylation; protein processing

Correspondence

K. Gevaert, Department of Biochemistry,

Faculty of Medicine and Health Sciences,

Ghent University, A. Baertsoenkaai 3,

B-9000 Ghent, Belgium

Fax: +32 92649496

Tel: +32 92649274

E-mail: [email protected]

Website: http://www.proteomics.be

(Received 24 April 2007, revised 10 Septem￾ber 2007, accepted 17 October 2007)

doi:10.1111/j.1742-4658.2007.06149.x

Numerous gel-free proteomics techniques have been reported over the past

few years, introducing a move from proteins to peptides as bits of informa￾tion in qualitative and quantitative proteome studies. Many shotgun pro￾teomics techniques randomly sample thousands of peptides in a qualitative

and quantitative manner but overlook the vast majority of protein modifi￾cations that are often crucial for proper protein structure and function.

Peptide-based proteomic approaches have thus been developed to profile a

diverse set of modifications including, but not at all limited, to phosphory￾lation, glycosylation and ubiquitination. Typical here is that each modifica￾tion needs a specific, tailor-made analytical procedure. In this minireview,

we discuss how one technique ) diagonal reverse-phase chromatogra￾phy ) is applied to study two different types of protein modification: pro￾tein processing and protein N-glycosylation. Additionally, we discuss an

activity-based proteome study in which purine-binding proteins were pro￾filed by diagonal chromatography.

Abbreviations

ABP, activity-based probe; COFRADIC, combined fractional diagonal chromatography; FSBA, 5¢-p-fluorosulfonylbenzoyladenosine; FSBG,

5¢-p-fluorosulfonylbenzoylguanosine; iTRAQ, isobaric tags for relative and absolute quantification; MudPIT, multidimensional protein

identification technology; PNGaseF, peptide N-glycosidase F; SB, sulfobenzoyl; SILAC, stable isotope labeling by amino acids in cell culture;

Nbs2, 2,4,6-trinitrobenzenesulfonic acid.

FEBS Journal 274 (2007) 6277–6289 ª 2007 The Authors Journal compilation ª 2007 FEBS 6277