Details of experiments between O95685 and P62258
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: PPP1R3D
Protein A fragment: 69-78
Protein A site: 14-3-3_pSer74
Protein A modification: (SEP)74
Protein A sequence: LRRARSLPSS
Protein A construct: LRRAR(pSer)LPSS (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: 0.96
Holdup BI standard deviation: 0.01
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): 6.34
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: 5.2
Experiment series 2
Protein A protein: PPP1R3D
Protein A fragment: 69-78
Protein A site: 14-3-3_pSer74
Protein A modification: (SEP)74
Protein A sequence: LRRARSLPSS
Protein A construct: LRRAR(pSer)LPSS (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: 0.95
Holdup BI standard deviation: 0.01
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): 5.69
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: 5.26
Experiment series 3
Protein A protein: PPP1R3D
Protein A fragment: 72-81
Protein A site: 14-3-3_pSer77
Protein A modification: (SEP)77
Protein A sequence: ARSLPSSPER
Protein A construct: ARSLP(pSer)SPER (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: 0.05
Holdup BI standard deviation: 0.1
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): 0.51
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.
Experiment series 4
Protein A protein: PPP1R3D
Protein A fragment: 72-81
Protein A site: 14-3-3_pSer77
Protein A modification: (SEP)77
Protein A sequence: ARSLPSSPER
Protein A construct: ARSLP(pSer)SPER (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: 0.03
Holdup BI standard deviation: 0.09
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): 0.31
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
The affinity was below the detection threshold of the assay.
Experiment series 5
Protein A protein: PPP1R3D
Protein A fragment: 20-29
Protein A site: 14-3-3_pSer25
Protein A modification: (SEP)25
Protein A sequence: GPRSLSCLSD
Protein A construct: GPRSL(pSer)CLSD (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: -0.01
Holdup BI standard deviation: 0.89
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): 0.01
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.