Probing SH2-domains using Inhibitor Affinity Purification (IAP)

Höfener M, Heinzlmeir S, Kuster B, Sewald N (2014)
Proteome Science 12(1).

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Abstract
Background Many human diseases are correlated with the dysregulation of signal transduction processes. One of the most important protein interaction domains in the context of signal transduction is the Src homology 2 (SH2) domain that binds phosphotyrosine residues. Hence, appropriate methods for the investigation of SH2 proteins are indispensable in diagnostics and medicinal chemistry. Therefore, an affinity resin for the enrichment of all SH2 proteins in one experiment would be desirable. However, current methods are unable to address all SH2 proteins simultaneously with a single compound or a small array of compounds. Results In order to overcome these limitations for the investigation of this particular protein family in future experiments, a dipeptide-derived probe has been designed, synthesized and evaluated. This probe successfully enriched 22 SH2 proteins from mixed cell lysates which contained 50 SH2 proteins. Further characterization of the SH2 binding properties of the probe using depletion and competition experiments indicated its ability to enrich complexes consisting of SH2 domain bearing regulatory PI3K subunits and catalytic phosphoinositide 3-kinase (PI3K) subunits that have no SH2 domain. Conclusion The results make this probe a promising starting point for the development of a mixed affinity resin with complete SH2 protein coverage. Moreover, the additional findings render it a valuable tool for the evaluation of PI3K complex interrupting inhibitors.
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Article Processing Charge funded by the Deutsche Forschungsgemeinschaft and the Open Access Publication Fund of Bielefeld University.
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Höfener M, Heinzlmeir S, Kuster B, Sewald N. Probing SH2-domains using Inhibitor Affinity Purification (IAP). Proteome Science. 2014;12(1).
Höfener, M., Heinzlmeir, S., Kuster, B., & Sewald, N. (2014). Probing SH2-domains using Inhibitor Affinity Purification (IAP). Proteome Science, 12(1).
Höfener, M., Heinzlmeir, S., Kuster, B., and Sewald, N. (2014). Probing SH2-domains using Inhibitor Affinity Purification (IAP). Proteome Science 12.
Höfener, M., et al., 2014. Probing SH2-domains using Inhibitor Affinity Purification (IAP). Proteome Science, 12(1).
M. Höfener, et al., “Probing SH2-domains using Inhibitor Affinity Purification (IAP)”, Proteome Science, vol. 12, 2014.
Höfener, M., Heinzlmeir, S., Kuster, B., Sewald, N.: Probing SH2-domains using Inhibitor Affinity Purification (IAP). Proteome Science. 12, (2014).
Höfener, Michael, Heinzlmeir, Stephanie, Kuster, Bernhard, and Sewald, Norbert. “Probing SH2-domains using Inhibitor Affinity Purification (IAP)”. Proteome Science 12.1 (2014).
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