Details of experiments between the fragments Q8TF72~1216-1225 and P62258~1-255
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: SHROOM3
Protein A fragment: 1216-1225
Protein A site: 14-3-3_pSer1221
Protein A modification: (SEP)1221
Protein A sequence: AGKSMSAEDL
Protein A construct: AGKSM(pSer)AEDL (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.15
Holdup BI standard deviation: 0.06
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 2.7
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021.10.13.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 5))
Number of measurements: 5
Measured pKd: 3.08
Experiment series 2
Protein A protein: SHROOM3
Protein A fragment: 1216-1225
Protein A site: 14-3-3_pSer1221
Protein A modification: (SEP)1221
Protein A sequence: AGKSMSAEDL
Protein A construct: AGKSM(pSer)AEDL (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.26
Holdup BI standard deviation: 0.15
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 1.76
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
Measured pKd: 3.49