Details of experiments between P31946 and Q12778
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: FOXO1
Protein B fragment: 177-186
Protein B site: 14-3-3_pThr182
Protein B modification: (TPO)182
Protein B sequence: AEKRLTLSQI
Protein B construct: AEKRL(pThr)LSQI (crude synthetic peptide)
Average holdup BI: 0.19
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): 1.86
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 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: FOXO1
Protein B fragment: 271-280
Protein B site: 14-3-3_pSer276
Protein B modification: (SEP)276
Protein B sequence: AKKKASLQSG
Protein B construct: AKKKA(pSer)LQSG (crude synthetic peptide)
Average holdup BI: 0.66
Holdup BI standard deviation: 0.42
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): 1.65
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: 4.27
Experiment series 3
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: FOXO1
Protein B fragment: 213-222
Protein B site: 14-3-3_pSer218
Protein B modification: (SEP)218
Protein B sequence: IRHNLSLHSK
Protein B construct: IRHNL(pSer)LHSK (crude synthetic peptide)
Average holdup BI: 0.37
Holdup BI standard deviation: 0.25
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): 1.64
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: 3.76
Experiment series 4
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: FOXO1
Protein B fragment: 644-653
Protein B site: 14-3-3_pThr649
Protein B modification: (TPO)649
Protein B sequence: SVKTTTHSWV
Protein B construct: SVKTT(pThr)HSWV (crude synthetic peptide)
Average holdup BI: 0.08
Holdup BI standard deviation: 0.11
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): 0.81
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: 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: FOXO1
Protein B fragment: 177-186
Protein B site: 14-3-3_pThr182
Protein B modification: (TPO)182
Protein B sequence: AEKRLTLSQI
Protein B construct: AEKRL(pThr)LSQI (crude synthetic peptide)
Average holdup BI: 0.06
Holdup BI standard deviation: 0.09
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): 0.68
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 6
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: FOXO1
Protein B fragment: 320-329
Protein B site: 14-3-3_pSer325
Protein B modification: (SEP)325
Protein B sequence: NASTISGRLS
Protein B construct: NASTI(pSer)GRLS (crude synthetic peptide)
Average holdup BI: 0.02
Holdup BI standard deviation: 0.04
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): 0.64
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 7
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: FOXO1
Protein B fragment: 213-222
Protein B site: 14-3-3_pSer218
Protein B modification: (SEP)218
Protein B sequence: IRHNLSLHSK
Protein B construct: IRHNL(pSer)LHSK (crude synthetic peptide)
Average holdup BI: -0.45
Holdup BI standard deviation: 1.32
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): 0.3
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 8
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: FOXO1
Protein B fragment: 251-260
Protein B site: 14-3-3_pSer256
Protein B modification: (SEP)256
Protein B sequence: RRRAASMDNN
Protein B construct: RRRAA(pSer)MDNN (crude synthetic peptide)
Average holdup BI: -0.08
Holdup BI standard deviation: 0.31
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): 0.23
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 9
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: FOXO1
Protein B fragment: 271-280
Protein B site: 14-3-3_pSer276
Protein B modification: (SEP)276
Protein B sequence: AKKKASLQSG
Protein B construct: AKKKA(pSer)LQSG (crude synthetic peptide)
Average holdup BI: -0.27
Holdup BI standard deviation: 1.28
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): 0.2
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 10
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: FOXO1
Protein B fragment: 644-653
Protein B site: 14-3-3_pThr649
Protein B modification: (TPO)649
Protein B sequence: SVKTTTHSWV
Protein B construct: SVKTT(pThr)HSWV (crude synthetic peptide)
Average holdup BI: 0.1
Holdup BI standard deviation: 0.56
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): 0.11
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 11
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: FOXO1
Protein B fragment: 320-329
Protein B site: 14-3-3_pSer325
Protein B modification: (SEP)325
Protein B sequence: NASTISGRLS
Protein B construct: NASTI(pSer)GRLS (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.04
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): 0.08
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 12
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: FOXO1
Protein B fragment: 251-260
Protein B site: 14-3-3_pSer256
Protein B modification: (SEP)256
Protein B sequence: RRRAASMDNN
Protein B construct: RRRAA(pSer)MDNN (crude synthetic peptide)
Average holdup BI: -0.04
Holdup BI standard deviation: 0.86
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): 0.04
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 13
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: FOXO1
Protein B fragment: 317-326
Protein B site: 14-3-3_pSer322
Protein B modification: (SEP)322
Protein B sequence: TSSNASTISG
Protein B construct: TSSNA(pSer)TISG (crude synthetic peptide)
Average holdup BI: 0.02
Holdup BI standard deviation: 0.33
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): 0.03
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.