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Detection and identification of plasma proteins that bind GlialCAM using ProteinChip™ Arrays, SELDI‐TOF MS, and nano‐LC MS/MS
Author(s) -
FavreKontula Linda,
SattonnetRoche Pascale,
Magnenat Edith,
Proudfoot Amanda E. I.,
Boschert Ursula,
Xenarios Ioannis,
Vilbois Francis,
Antonsson Bruno
Publication year - 2008
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.200700564
Subject(s) - chemistry , glycoprotein , chromatography , mass spectrometry , proteome , computational biology , affinity chromatography , lectin , immunoprecipitation , transmembrane protein , biochemistry , biology , receptor , gene , enzyme
Abstract In order to fully understand biological processes it is essential to identify interactions in protein complexes. There are several techniques available to study this type of interactions, such as yeast two‐hybrid screens, affinity chromatography, and coimmunoprecipitation. We propose a novel strategy to identify protein–protein interactions, comprised of first detecting the interactions using ProteinChips and SELDI‐TOF MS, followed by the isolation of the interacting proteins through affinity beads and RP‐HPLC and finally identifying the proteins using nano‐LC MS/MS. The advantages of this new strategy are that the primary high‐throughput screening of samples can be performed with small amounts of sample, no specific antibody is needed and the proteins represented on the SELDI‐TOF MS spectra can be identified with high confidence. Furthermore, the method is faster and less labor‐intensive than other current approaches. Using this novel method, we isolated and identified the interactions of two mouse plasma proteins, mannose binding lectin C and properdin, with GlialCAM, a type 1 transmembrane glycoprotein that belongs to the Ig superfamily.