Details of experiments between P61981 and O60825
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: PFKFB2
Protein B fragment: 470-479
Protein B site: 14-3-3_pThr475
Protein B modification: (TPO)475
Protein B sequence: LSSSNTIRRP
Protein B construct: LSSSN(pThr)IRRP (crude synthetic peptide)
Average holdup BI: 0.66
Holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 129
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 7.21
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: 4.18
Experiment series 2
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: PFKFB2
Protein B fragment: 470-479
Protein B site: 14-3-3_pThr475
Protein B modification: (TPO)475
Protein B sequence: LSSSNTIRRP
Protein B construct: LSSSN(pThr)IRRP (crude synthetic peptide)
Average holdup BI: 0.74
Holdup BI standard deviation: 0.03
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): 5.73
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: 4.33
Experiment series 3
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: PFKFB2
Protein B fragment: 470-479
Protein B site: 14-3-3_pThr475
Protein B modification: (TPO)475
Protein B sequence: LSSSNTIRRP
Protein B construct: LSSSN(pThr)IRRP (crude synthetic peptide)
Average holdup BI: 0.68
Holdup BI standard deviation: 0.04
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): 4.82
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: 4.29
Experiment series 4
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: PFKFB2
Protein B fragment: 461-470
Protein B site: 14-3-3_pSer466
Protein B modification: (SEP)466
Protein B sequence: RMRRNSFTPL
Protein B construct: RMRRN(pSer)FTPL (crude synthetic peptide)
Average holdup BI: 0.56
Holdup BI standard deviation: 0.08
Immobilized partner concentration in holdup experiment (10-6M): 129
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 4.28
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
Experiment series 5
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: PFKFB2
Protein B fragment: 478-487
Protein B site: 14-3-3_pSer483
Protein B modification: (SEP)483
Protein B sequence: RPRNYSVGSR
Protein B construct: RPRNY(pSer)VGSR (crude synthetic peptide)
Average holdup BI: 0.65
Holdup BI standard deviation: 0.13
Immobilized partner concentration in holdup experiment (10-6M): 129
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 4.27
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: 4.16
Experiment series 6
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: PFKFB2
Protein B fragment: 478-487
Protein B site: 14-3-3_pSer483
Protein B modification: (SEP)483
Protein B sequence: RPRNYSVGSR
Protein B construct: RPRNY(pSer)VGSR (crude synthetic peptide)
Average holdup BI: 0.58
Holdup BI standard deviation: 0.13
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): 3.82
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: 4.09
Experiment series 7
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: PFKFB2
Protein B fragment: 478-487
Protein B site: 14-3-3_pSer483
Protein B modification: (SEP)483
Protein B sequence: RPRNYSVGSR
Protein B construct: RPRNY(pSer)VGSR (crude synthetic peptide)
Average holdup BI: 0.45
Holdup BI standard deviation: 0.11
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): 3.03
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: 3.78
Experiment series 8
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: PFKFB2
Protein B fragment: 461-470
Protein B site: 14-3-3_pSer466
Protein B modification: (SEP)466
Protein B sequence: RMRRNSFTPL
Protein B construct: RMRRN(pSer)FTPL (crude synthetic peptide)
Average holdup BI: 0.45
Holdup BI standard deviation: 0.15
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): 1.93
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: 3.78
Experiment series 9
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: PFKFB2
Protein B fragment: 461-470
Protein B site: 14-3-3_pSer466
Protein B modification: (SEP)466
Protein B sequence: RMRRNSFTPL
Protein B construct: RMRRN(pSer)FTPL (crude synthetic peptide)
Average holdup BI: 0.58
Holdup BI standard deviation: 0.2
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): 1.34
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: 4.1