Details of experiments between Q96Q42 and Q04917
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: ALS2
Protein A fragment: 461-470
Protein A site: 14-3-3_pSer466
Protein A modification: (SEP)466
Protein A sequence: GKKSSSLVDI
Protein A construct: GKKSS(pSer)LVDI (crude synthetic peptide)
Protein B protein: YWHAH
Protein B fragment: 1-246
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3eta
Average holdup BI: 0.53
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 6.67
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: 3.94
Experiment series 2
Protein A protein: ALS2
Protein A fragment: 525-534
Protein A site: 14-3-3_pThr530
Protein A modification: (TPO)530
Protein A sequence: RTEVWTWGKG
Protein A construct: RTEVW(pThr)WGKG (crude synthetic peptide)
Protein B protein: YWHAH
Protein B fragment: 1-246
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3eta
Average holdup BI: 0.82
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 5.74
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: 4.53
Experiment series 3
Protein A protein: ALS2
Protein A fragment: 478-487
Protein A site: 14-3-3_pSer483
Protein A modification: (SEP)483
Protein A sequence: GSRRLSLPGL
Protein A construct: GSRRL(pSer)LPGL (crude synthetic peptide)
Protein B protein: YWHAH
Protein B fragment: 1-246
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3eta
Average holdup BI: 0.74
Holdup BI standard deviation: 0.06
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 4.25
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: 4.34