Mechanism of Beta-Arrestin 1 Mediated Src Activation via Src SH3 Domain
- Description
Beta-arrestins (βarrs) are key transducers of G-protein coupled receptors, but the mechanisms of βarr signaling are poorly understood. Therefore, this study examined the structural mechanisms underlying βarr-mediated signal transduction. Using cryo-electron microscopy (cryo-EM), the study examined how βarr1 recruits and activates the non-receptor tyrosine kinase Src, a well-established signaling partner of βarrs. To map the binding interface between the SH3 domain of Src and βarr1, a disulfide trapping strategy was utilized. The βarr1 and SH3 Src sequences were derived from rat (Rattus norvegicus) and chicken (Gallus gallus).
Since the SH3–βarr1 complexes were stabilized using cysteine substitutions and disulfide trapping, the interaction interfaces and dynamics of free βarr1, V2Rpp-activated βarr1, and SH3 were mapped using hydrogen–deuterium exchange mass spectrometry (HDX-MS) to determine whether these interactions also occur under physiological conditions, in the absence of engineered mutations. To gain insight into how inactive βarr1 engages SH1, lysine-specific cross-linking mass spectrometry (CXMS) was performed using purified βarr1 and SH1. This dataset contains cryo-EM maps of SH3–βarr1-CC complex, SH3–βarr1-N complex, and Src–βarr1-CC complex, and atomic coordinates of SH3–βarr1-CC complex, SH3–βarr1-N complex, and Src–βarr1-CC complex. In addition, the dataset includes HDX-MS and CXMS data.
Access
- Restrictions
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Free to All
- Instructions
- The cryo-EM maps have been deposited in the Electron Microscopy Data Bank (EMDB) and the atomic coordinates have been deposited in the Protein Data Bank (PDB). The HDX-MS data have been deposited to the Mass Spectrometry Interactive Virtual Environment (MassIVE) repository and the CXMS data have been deposited to the PRoteomics IDEntifications (PRIDE) database. Supplementary and source data are provided within the article on PubMed Central (PMC).
- Grant Support
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LT000174/2018/Human Frontier Science ProgramALTF 1071-2017/European Molecular Biology Organization45126409/The Pittsburgh Foundation2023CHAL4223/Prostate Cancer FoundationMoonshot Biomarker Program/Allegheny Health Network Cancer Institute and Highmark Health4100095527/Pennsylvania State Formula Grant