Details of experiments between P31946 and O43524
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: FOXO3
Protein B fragment: 408-417
Protein B site: 14-3-3_pSer413
Protein B modification: (SEP)413
Protein B sequence: MQRSSSFPYT
Protein B construct: MQRSS(pSer)FPYT (crude synthetic peptide)
Average holdup BI: 0.9
Holdup BI standard deviation: 0.01
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): 7.97
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: 4.96
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: FOXO3
Protein B fragment: 408-417
Protein B site: 14-3-3_pSer413
Protein B modification: (SEP)413
Protein B sequence: MQRSSSFPYT
Protein B construct: MQRSS(pSer)FPYT (crude synthetic peptide)
Average holdup BI: 0.95
Holdup BI standard deviation: 0.01
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): 5.83
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: 5.14
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: FOXO3
Protein B fragment: 550-559
Protein B site: 14-3-3_pSer555
Protein B modification: (SEP)555
Protein B sequence: LSNSVSNMGL
Protein B construct: LSNSV(pSer)NMGL (crude synthetic peptide)
Average holdup BI: 0.35
Holdup BI standard deviation: 0.1
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): 2.49
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: 3.61
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: FOXO3
Protein B fragment: 550-559
Protein B site: 14-3-3_pSer555
Protein B modification: (SEP)555
Protein B sequence: LSNSVSNMGL
Protein B construct: LSNSV(pSer)NMGL (crude synthetic peptide)
Average holdup BI: 0.24
Holdup BI standard deviation: 0.19
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): 1.39
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: 3.5
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: FOXO3
Protein B fragment: 174-183
Protein B site: 14-3-3_pThr179
Protein B modification: (TPO)179
Protein B sequence: PDKRLTLSQI
Protein B construct: PDKRL(pThr)LSQI (crude synthetic peptide)
Average holdup BI: 0.13
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): 0.89
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 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: FOXO3
Protein B fragment: 275-284
Protein B site: 14-3-3_pSer280
Protein B modification: (SEP)280
Protein B sequence: QTAPESADDS
Protein B construct: QTAPE(pSer)ADDS (crude synthetic peptide)
Average holdup BI: -0.47
Holdup BI standard deviation: 0.84
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): 0.8
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 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: FOXO3
Protein B fragment: 316-325
Protein B site: 14-3-3_pSer321
Protein B modification: (SEP)321
Protein B sequence: NASTVSGRLS
Protein B construct: NASTV(pSer)GRLS (crude synthetic peptide)
Average holdup BI: 0.03
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): 0.8
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 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: FOXO3
Protein B fragment: 204-213
Protein B site: 14-3-3_pSer209
Protein B modification: (SEP)209
Protein B sequence: AGWKNSIRHN
Protein B construct: AGWKN(pSer)IRHN (crude synthetic peptide)
Average holdup BI: -0.07
Holdup BI standard deviation: 0.18
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): 0.31
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: FOXO3
Protein B fragment: 394-403
Protein B site: 14-3-3_pSer399
Protein B modification: (SEP)399
Protein B sequence: ITLPPSQPSP
Protein B construct: ITLPP(pSer)QPSP (crude synthetic peptide)
Average holdup BI: -0.48
Holdup BI standard deviation: 1.69
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): 0.26
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
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: FOXO3
Protein B fragment: 174-183
Protein B site: 14-3-3_pThr179
Protein B modification: (TPO)179
Protein B sequence: PDKRLTLSQI
Protein B construct: PDKRL(pThr)LSQI (crude synthetic peptide)
Average holdup BI: 0.05
Holdup BI standard deviation: 0.15
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): 0.26
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 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: FOXO3
Protein B fragment: 210-219
Protein B site: 14-3-3_pSer215
Protein B modification: (SEP)215
Protein B sequence: IRHNLSLHSR
Protein B construct: IRHNL(pSer)LHSR (crude synthetic peptide)
Average holdup BI: -0.03
Holdup BI standard deviation: 0.12
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 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: FOXO3
Protein B fragment: 316-325
Protein B site: 14-3-3_pSer321
Protein B modification: (SEP)321
Protein B sequence: NASTVSGRLS
Protein B construct: NASTV(pSer)GRLS (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.02
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 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: FOXO3
Protein B fragment: 204-213
Protein B site: 14-3-3_pSer209
Protein B modification: (SEP)209
Protein B sequence: AGWKNSIRHN
Protein B construct: AGWKN(pSer)IRHN (crude synthetic peptide)
Average holdup BI: 0.03
Holdup BI standard deviation: 0.24
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): 0.11
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 14
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: FOXO3
Protein B fragment: 583-592
Protein B site: 14-3-3_pSer588
Protein B modification: (SEP)588
Protein B sequence: LSDSLSGSSL
Protein B construct: LSDSL(pSer)GSSL (crude synthetic peptide)
Average holdup BI: -0.1
Holdup BI standard deviation: 1
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 15
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: FOXO3
Protein B fragment: 227-236
Protein B site: 14-3-3_pSer232
Protein B modification: (SEP)232
Protein B sequence: GTGKSSWWII
Protein B construct: GTGKS(pSer)WWII (crude synthetic peptide)
Average holdup BI: -0.02
Holdup BI standard deviation: 0.32
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): 0.05
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
The affinity was below the detection threshold of the assay.
Experiment series 16
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: FOXO3
Protein B fragment: 210-219
Protein B site: 14-3-3_pSer215
Protein B modification: (SEP)215
Protein B sequence: IRHNLSLHSR
Protein B construct: IRHNL(pSer)LHSR (crude synthetic peptide)
Average holdup BI: -0.01
Holdup BI standard deviation: 0.32
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): 0.02
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.