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Proteomics. 2015 Apr;15(7):1230-4. doi: 10.1002/pmic.201400395. Epub 2015 Feb 3.

Sortase A mediated site-specific immobilization for identification of protein interactions in affinity purification-mass spectrometry experiments.

Author information

1
Leibniz-Institut für Molekulare Pharmakologie, Berlin, Germany; Institut für Chemie und Biochemie, Freie Universität Berlin, Berlin, Germany.

Abstract

Proteomics approaches using MS in combination with affinity purification have emerged as powerful tools to study protein-protein interactions. Here we make use of the specificity of sortase A transpeptidation reaction to prepare affinity matrices in which a protein bait is covalently linked to the matrix via a short C-terminal linker region. As a result of this site-directed immobilization, the bait remains functionally accessible to protein interactions. To apply this approach, we performed SILAC-based pull-down experiments and demonstrate the suitability of the approach.

KEYWORDS:

Mass spectrometry; Protein interactions; SILAC; Sortase A; Technology

PMID:
25504886
DOI:
10.1002/pmic.201400395
[Indexed for MEDLINE]

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