Details of experiments between P31947 and O95208
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 267-276
Protein B site: 14-3-3_pThr272
Protein B modification: (TPO)272
Protein B sequence: QARPQTSGEE
Protein B construct: QARPQ(pThr)SGEE (crude synthetic peptide)
Measured dissociation constant (10-6M): <100
Immobilized partner concentration in holdup experiment (10-6M): 113
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: 1
Measured pKd: 4
Experiment series 2
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 190-199
Protein B site: 14-3-3_pSer195
Protein B modification: (SEP)195
Protein B sequence: GGSPASYHGS
Protein B construct: GGSPA(pSer)YHGS (crude synthetic peptide)
Average holdup BI: 0.32
Holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 142
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 5.98
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: 3.52
Experiment series 3
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 168-177
Protein B site: 14-3-3_pSer173
Protein B modification: (SEP)173
Protein B sequence: FGRGSSQPNL
Protein B construct: FGRGS(pSer)QPNL (crude synthetic peptide)
Average holdup BI: 0.37
Holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 142
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 5.91
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: 3.62
Experiment series 4
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 168-177
Protein B site: 14-3-3_pSer173
Protein B modification: (SEP)173
Protein B sequence: FGRGSSQPNL
Protein B construct: FGRGS(pSer)QPNL (crude synthetic peptide)
Average holdup BI: 0.31
Holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 113
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 4.67
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: 3.6
Experiment series 5
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A modification: 100_uM_FCA
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 168-177
Protein B site: 14-3-3_pSer173
Protein B modification: (SEP)173
Protein B sequence: FGRGSSQPNL
Protein B construct: FGRGS(pSer)QPNL (crude synthetic peptide)
Average holdup BI: 0.23
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 111
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 3.6
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: 3.42
Experiment series 6
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A modification: 100_uM_FCA
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 190-199
Protein B site: 14-3-3_pSer195
Protein B modification: (SEP)195
Protein B sequence: GGSPASYHGS
Protein B construct: GGSPA(pSer)YHGS (crude synthetic peptide)
Average holdup BI: 0.23
Holdup BI standard deviation: 0.08
Immobilized partner concentration in holdup experiment (10-6M): 111
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 2.45
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: 3.42
Experiment series 7
Protein A protein: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 190-199
Protein B site: 14-3-3_pSer195
Protein B modification: (SEP)195
Protein B sequence: GGSPASYHGS
Protein B construct: GGSPA(pSer)YHGS (crude synthetic peptide)
Average holdup BI: 0.19
Holdup BI standard deviation: 0.12
Immobilized partner concentration in holdup experiment (10-6M): 113
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 1.4
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A modification: 100_uM_FCA
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 322-331
Protein B site: 14-3-3_pSer327
Protein B modification: (SEP)327
Protein B sequence: KKEHGSLPQQ
Protein B construct: KKEHG(pSer)LPQQ (crude synthetic peptide)
Average holdup BI: -0.5
Holdup BI standard deviation: 0.89
Immobilized partner concentration in holdup experiment (10-6M): 111
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.66
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A modification: 100_uM_FCA
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 403-412
Protein B site: 14-3-3_pSer408
Protein B modification: (SEP)408
Protein B sequence: SVPKNSDPWA
Protein B construct: SVPKN(pSer)DPWA (crude synthetic peptide)
Average holdup BI: -0.05
Holdup BI standard deviation: 0.09
Immobilized partner concentration in holdup experiment (10-6M): 111
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.56
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 403-412
Protein B site: 14-3-3_pSer408
Protein B modification: (SEP)408
Protein B sequence: SVPKNSDPWA
Protein B construct: SVPKN(pSer)DPWA (crude synthetic peptide)
Average holdup BI: 0.09
Holdup BI standard deviation: 0.16
Immobilized partner concentration in holdup experiment (10-6M): 113
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.47
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A modification: 100_uM_FCA
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 311-320
Protein B site: 14-3-3_pThr316
Protein B modification: (TPO)316
Protein B sequence: ESRRDTVKIP
Protein B construct: ESRRD(pThr)VKIP (crude synthetic peptide)
Average holdup BI: 0.05
Holdup BI standard deviation: 0.11
Immobilized partner concentration in holdup experiment (10-6M): 111
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.4
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 322-331
Protein B site: 14-3-3_pSer327
Protein B modification: (SEP)327
Protein B sequence: KKEHGSLPQQ
Protein B construct: KKEHG(pSer)LPQQ (crude synthetic peptide)
Average holdup BI: 0.13
Holdup BI standard deviation: 0.43
Immobilized partner concentration in holdup experiment (10-6M): 142
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.24
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 403-412
Protein B site: 14-3-3_pSer408
Protein B modification: (SEP)408
Protein B sequence: SVPKNSDPWA
Protein B construct: SVPKN(pSer)DPWA (crude synthetic peptide)
Average holdup BI: 0.02
Holdup BI standard deviation: 0.09
Immobilized partner concentration in holdup experiment (10-6M): 142
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.17
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 322-331
Protein B site: 14-3-3_pSer327
Protein B modification: (SEP)327
Protein B sequence: KKEHGSLPQQ
Protein B construct: KKEHG(pSer)LPQQ (crude synthetic peptide)
Average holdup BI: 0.04
Holdup BI standard deviation: 0.22
Immobilized partner concentration in holdup experiment (10-6M): 113
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.13
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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 311-320
Protein B site: 14-3-3_pThr316
Protein B modification: (TPO)316
Protein B sequence: ESRRDTVKIP
Protein B construct: ESRRD(pThr)VKIP (crude synthetic peptide)
Average holdup BI: 0.01
Holdup BI standard deviation: 0.07
Immobilized partner concentration in holdup experiment (10-6M): 142
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: 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: SFN
Protein A fragment: 1-248
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3sigma
Protein B protein: EPN2
Protein B fragment: 311-320
Protein B site: 14-3-3_pThr316
Protein B modification: (TPO)316
Protein B sequence: ESRRDTVKIP
Protein B construct: ESRRD(pThr)VKIP (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.07
Immobilized partner concentration in holdup experiment (10-6M): 113
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: 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.