Quantifying G Protein Subunits in Co-IP

Co-IP samples containing Gβ and Gγ subunits were separated, digested, and analyzed by a TSQ Vantage triple quadrupole mass spectrometer (Thermo Scientific)^{28 (link)}. To allow comparisons between G proteins co-IPed from multiple mice, quantitative Gβ and Gγ subunits detected (fmol) were normalized by the amount of protein (mg) used in co-IPs. The amount of protein used in co-IPs was calculated using the volume of precleared lysate used and the protein concentration of precleared lysate from BCA assay.

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Yim Y.Y., Betke K.M., McDonald W.H., Gilsbach R., Chen Y., Hyde K., Wang Q., Hein L, & Hamm H. (2019). The in vivo specificity of synaptic Gβ and Gγ subunits to the α2a adrenergic receptor at CNS synapses. Scientific Reports, 9, 1718.

Publication 2019

TSQ Vantage

Assay G proteins Mice Protein Subunits

Corresponding Organization :

Other organizations : Vanderbilt University, University of Freiburg, University of Alabama at Birmingham

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Quantitative amount of Gβ and Gγ subunits (fmol) detected by mass spectrometry

control variables

Amount of protein (mg) used in co-immunoprecipitation (co-IP) experiments, calculated from the volume of precleared lysate used and the protein concentration of precleared lysate from BCA assay

controls

Positive control: None mentioned
Negative control: None mentioned

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