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High Throughput Affinity Purification and Mass Spectrometry to Determine Protein Complex Interactions

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New Frontiers of Network Analysis in Systems Biology

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Abstract

Affinity purification, or immunoprecipitation, followed by mass spectrometry (AP/MS or IP/MS, respectively) is a proper and powerful method to analyze protein complexes. Technological capabilities of mass spectrometry instrumentation have dramatically improved over the last decade, and now allow routine detection of high- and low-abundance components in composite protein mixtures, expanding our ability to use co-immunoprecipitation to study cellular regulatory protein interaction networks. In this review, we summarize several key efforts and accomplishments in applying mass spectrometry towards mapping of a proteome-wide interactome. Emerging high-throughput studies begin to etch a ‘systems biology’ view of protein-protein interactions, where modularity and interconnectivity of cellular protein machinery are unequivocally revealed. Yet, major challenges still remain in the application of high-throughput (HT) affinity purification for studies in higher organisms, particularly in human cells, and in addressing persistent issues associated with analysis, interpretation, and dissemination of massive IP/MS data. We describe some of the conceptual and practical challenges of the HT-IP/MS approach, and discuss feasible solutions in context of the protein complexes involved in transcriptional regulation.

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Acknowledgements

We thank Dr. Yaroslava Bulynko for critical reading of the manuscript. This work was supported by the NIH grants U19-DK62434 (including the Proteomics Strand funding to B.W.O. and J.Q. and the Collaborative Bridging Project funding to R.B.L.), NIDDK DK59820 (to B.W.O.), and NCI CA84199 (to J.Q.). We also acknowledge the support of the McLean Foundation through the Center for Molecular Discovery at Baylor College of Medicine.

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  1. Baylor College of Medicine, One Baylor Plaza, BCM 130, Houston, TX, 77030, USA

    Anna Malovannaya, Rainer B. Lanz, Bert W. O’Malley & Jun Qin

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  1. Anna Malovannaya
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  2. Rainer B. Lanz
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Correspondence to Anna Malovannaya .

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  1. Pharmacology & Systems Therapeutics, Systems Biology Center New York, Mount Sinai School of Medicine, New York, 10029, New York, USA

    Avi Ma'ayan

  2. Institute For Life Sciences, Southampton University, Highfield Campus, Southampton, SO17 1BJ, United Kingdom

    Ben D. MacArthur

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Malovannaya, A., Lanz, R.B., O’Malley, B.W., Qin, J. (2012). High Throughput Affinity Purification and Mass Spectrometry to Determine Protein Complex Interactions. In: Ma'ayan, A., MacArthur, B. (eds) New Frontiers of Network Analysis in Systems Biology. Springer, Dordrecht. https://doi.org/10.1007/978-94-007-4330-4_8

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