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Differential binding studies applying functional protein microarrays and surface plasmon resonance
Author(s) -
Seitz Harald,
Hutschenreiter Silke,
Hultschig Claus,
Zeilinger Carsten,
Zimmermann Bastian,
Kleinjung Frank,
Schuchhardt Johannes,
Eickhoff Holger,
Herberg Friedrich W.
Publication year - 2006
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200500181
Subject(s) - protein array analysis , protein microarray , surface plasmon resonance , proteome , protein–protein interaction , computational biology , proteomics , dna microarray , chemistry , biophysics , biology , microbiology and biotechnology , biochemistry , nanotechnology , gene expression , materials science , gene , nanoparticle
A variety of different in vivo and in vitro technologies provide comprehensive insights in protein‐protein interaction networks. Here we demonstrate a novel approach to analyze, verify and quantify putative interactions between two members of the S100 protein family and 80 recombinant proteins derived from a proteome‐wide protein expression library. Surface plasmon resonance (SPR) using Biacore technology and functional protein microarrays were used as two independent methods to study protein‐protein interactions. With this combined approach we were able to detect nine calcium‐dependent interactions between Arg‐Gly‐Ser‐(RGS)‐His 6 tagged proteins derived from the library and GST‐tagged S100B and S100A6, respectively. For the protein microarray affinity‐purified proteins from the expression library were spotted onto modified glass slides and probed with the S100 proteins. SPR experiments were performed in the same setup and in a vice‐versa approach reversing analytes and ligands to determine distinct association and dissociation patterns of each positive interaction. Besides already known interaction partners, several novel binders were found independently with both detection methods, albeit analogous immobilization strategies had to be applied in both assays.