Details of experiments between the fragments P61981~1-247 and Q5T5U3~5-14
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: YWHAG
Protein A fragment: 1-247
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3gamma
Protein B protein: ARHGAP21
Protein B fragment: 5-14
Protein B site: 14-3-3_pSer10
Protein B modification: (SEP)10
Protein B sequence: RRTGLSEGDG
Protein B construct: RRTGL(pSer)EGDG (crude synthetic peptide)
Measured dissociation constant (10-6M): <100
Immobilized partner concentration in holdup experiment (10-6M): 134
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.02.17.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 4))
Number of measurements: 1
Measured pKd: 4
Experiment series 2
Protein A protein: YWHAG
Protein A fragment: 1-247
Protein A site: 14-3-3
Protein A modification: 100_uM_FCA
Protein A construct: avi-his6-MBP-TEVsite-14-3-3gamma
Protein B protein: ARHGAP21
Protein B fragment: 5-14
Protein B site: 14-3-3_pSer10
Protein B modification: (SEP)10
Protein B sequence: RRTGLSEGDG
Protein B construct: RRTGL(pSer)EGDG (crude synthetic peptide)
Average holdup BI: -0.07
Holdup BI standard deviation: 0.64
Immobilized partner concentration in holdup experiment (10-6M): 115
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.06
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.02.17.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 4))
Number of measurements: 4
The affinity was below the detection threshold of the assay.