Details of experiments between the fragments P31946~1-246 and O95208~190-199
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: YWHAB
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3beta
Protein B protein: EPN2
Protein B fragment: 190-199
Protein B site: 14-3-3_pSer195
Protein B modification: (SEP)195
Protein B sequence: GGSPASYHGS
Protein B construct: GGSPA(pSer)YHGS (crude synthetic peptide)
Average holdup BI: 0.54
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 102
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 8.05
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: 4.07
Experiment series 2
Protein A protein: YWHAB
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3beta
Protein B protein: EPN2
Protein B fragment: 190-199
Protein B site: 14-3-3_pSer195
Protein B modification: (SEP)195
Protein B sequence: GGSPASYHGS
Protein B construct: GGSPA(pSer)YHGS (crude synthetic peptide)
Average holdup BI: 0.47
Holdup BI standard deviation: 0.08
Immobilized partner concentration in holdup experiment (10-6M): 134
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 3.26
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: 3.81