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The challenge to quantify proteins with charge trains due to isoforms or conformers
Author(s) -
Deng Xi,
Hahne Thomas,
Schröder Simone,
Redweik Sabine,
Nebija Dashnor,
Schmidt Hendrik,
Janssen Ottmar,
Lachmann Bodo,
Wätzig Hermann
Publication year - 2012
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201100321
Subject(s) - train , software , isoelectric point , isoelectric focusing , charge (physics) , spots , gene isoform , glycosylation , software package , computer science , biological system , chemistry , computational biology , biology , biochemistry , physics , enzyme , gene , cartography , quantum mechanics , geography , programming language
Single proteins separated by 2‐DE often show multiple spots spreading along the first dimension. In many cases, such charge trains are explained by isoform differences or by putative post‐translational modifications including phosphorylation, glycosylation and others. We now report that individual spots of such charge trains on 2‐D gels in fact often represent the same protein, but, apparently due to conformational changes, segregate to different isoelectric points. If MS analysis reveals protein identity, we therefore suggest integrating all individual spots within a charge train for quantification. Especially in quality control of pharmaceutical proteins, the integration of the spot groups of all active contents is preferable in order to obtain reproducible and reasonable quantitative results. However, most commercial software packages for gel analysis integrate the signals spot‐wise. We provide an improved quantification tool for proteins with charge train groups. This calculation can be implemented using the MATLAB software and the self‐developed “Correct Integration Software System” or the commercial software package Delta2D.