Details of experiments between P61981 and Q9Y2J2
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: EPB41L3
Protein B fragment: 438-447
Protein B site: 14-3-3_pSer443
Protein B modification: (SEP)443
Protein B sequence: YTMSRSLDGA
Protein B construct: YTMSR(pSer)LDGA (crude synthetic peptide)
Average holdup BI: 0.85
Holdup BI standard deviation: 0.02
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): 9
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: 4.65
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: EPB41L3
Protein B fragment: 396-405
Protein B site: 14-3-3_pThr401
Protein B modification: (TPO)401
Protein B sequence: PKKFLTLGSK
Protein B construct: PKKFL(pThr)LGSK (crude synthetic peptide)
Average holdup BI: 0.87
Holdup BI standard deviation: 0.01
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): 8.56
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: 4.73
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: EPB41L3
Protein B fragment: 438-447
Protein B site: 14-3-3_pSer443
Protein B modification: (SEP)443
Protein B sequence: YTMSRSLDGA
Protein B construct: YTMSR(pSer)LDGA (crude synthetic peptide)
Average holdup BI: 0.77
Holdup BI standard deviation: 0.03
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): 7.2
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
Measured pKd: 4.48
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: EPB41L3
Protein B fragment: 438-447
Protein B site: 14-3-3_pSer443
Protein B modification: (SEP)443
Protein B sequence: YTMSRSLDGA
Protein B construct: YTMSR(pSer)LDGA (crude synthetic peptide)
Average holdup BI: 0.83
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): 6.11
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
Measured pKd: 4.56
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: EPB41L3
Protein B fragment: 396-405
Protein B site: 14-3-3_pThr401
Protein B modification: (TPO)401
Protein B sequence: PKKFLTLGSK
Protein B construct: PKKFL(pThr)LGSK (crude synthetic peptide)
Average holdup BI: 0.83
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): 6.05
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: 4.56
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: EPB41L3
Protein B fragment: 396-405
Protein B site: 14-3-3_pThr401
Protein B modification: (TPO)401
Protein B sequence: PKKFLTLGSK
Protein B construct: PKKFL(pThr)LGSK (crude synthetic peptide)
Average holdup BI: 0.87
Holdup BI standard deviation: 0.01
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): 5.87
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: 4.78
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: EPB41L3
Protein B fragment: 415-424
Protein B site: 14-3-3_pSer420
Protein B modification: (SEP)420
Protein B sequence: QTRRASALID
Protein B construct: QTRRA(pSer)ALID (crude synthetic peptide)
Average holdup BI: 0.11
Holdup BI standard deviation: 0.03
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): 3.83
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: 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: EPB41L3
Protein B fragment: 415-424
Protein B site: 14-3-3_pSer420
Protein B modification: (SEP)420
Protein B sequence: QTRRASALID
Protein B construct: QTRRA(pSer)ALID (crude synthetic peptide)
Average holdup BI: 0.11
Holdup BI standard deviation: 0.03
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): 3.42
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: 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: EPB41L3
Protein B fragment: 952-961
Protein B site: 14-3-3_pSer957
Protein B modification: (SEP)957
Protein B sequence: KTETISFGSV
Protein B construct: KTETI(pSer)FGSV (crude synthetic peptide)
Average holdup BI: 0.2
Holdup BI standard deviation: 0.07
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): 3.09
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 10
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: EPB41L3
Protein B fragment: 952-961
Protein B site: 14-3-3_pSer957
Protein B modification: (SEP)957
Protein B sequence: KTETISFGSV
Protein B construct: KTETI(pSer)FGSV (crude synthetic peptide)
Average holdup BI: 0.18
Holdup BI standard deviation: 0.07
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): 2.4
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 11
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: EPB41L3
Protein B fragment: 455-464
Protein B site: 14-3-3_pSer460
Protein B modification: (SEP)460
Protein B sequence: TTKGISQTNL
Protein B construct: TTKGI(pSer)QTNL (crude synthetic peptide)
Average holdup BI: 0.06
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): 1.62
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 12
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: EPB41L3
Protein B fragment: 415-424
Protein B site: 14-3-3_pSer420
Protein B modification: (SEP)420
Protein B sequence: QTRRASALID
Protein B construct: QTRRA(pSer)ALID (crude synthetic peptide)
Average holdup BI: 0.13
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): 1.17
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: 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: EPB41L3
Protein B fragment: 455-464
Protein B site: 14-3-3_pSer460
Protein B modification: (SEP)460
Protein B sequence: TTKGISQTNL
Protein B construct: TTKGI(pSer)QTNL (crude synthetic peptide)
Average holdup BI: 0.03
Holdup BI standard deviation: 0.03
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): 0.97
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 14
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: EPB41L3
Protein B fragment: 455-464
Protein B site: 14-3-3_pSer460
Protein B modification: (SEP)460
Protein B sequence: TTKGISQTNL
Protein B construct: TTKGI(pSer)QTNL (crude synthetic peptide)
Average holdup BI: 0.03
Holdup BI standard deviation: 0.05
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): 0.54
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: 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: EPB41L3
Protein B fragment: 756-765
Protein B site: 14-3-3_pThr761
Protein B modification: (TPO)761
Protein B sequence: EKRLSTSPVR
Protein B construct: EKRLS(pThr)SPVR (crude synthetic peptide)
Average holdup BI: 0.27
Holdup BI standard deviation: 0.77
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): 0.27
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
The affinity was below the detection threshold of the assay.
Experiment series 16
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: EPB41L3
Protein B fragment: 952-961
Protein B site: 14-3-3_pSer957
Protein B modification: (SEP)957
Protein B sequence: KTETISFGSV
Protein B construct: KTETI(pSer)FGSV (crude synthetic peptide)
Average holdup BI: 0.02
Holdup BI standard deviation: 0.06
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): 0.27
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 17
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: EPB41L3
Protein B fragment: 396-405
Protein B site: 14-3-3_pThr401
Protein B modification: (TPO)401
Protein B sequence: PKKFLTLGSK
Protein B construct: biotin-ttds-PKKFL(pThr)LGSK (HPLC-purified synthetic peptide)
Measured dissociation constant (10-6M): 7.9
Standard deviation of dissociation constant (10-6M): 0.81
Experiment method: competitive Fluorescence Polarization
Details of affinity fitting: Competitive binding equation in ProFit
Date of measurement: 2022.01.18
Number of measurements: 3
Measured pKd: 5.1
Experiment series 18
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: EPB41L3
Protein B fragment: 396-405
Protein B site: 14-3-3_pThr401
Protein B modification: (TPO)401
Protein B sequence: PKKFLTLGSK
Protein B construct: biotin-ttds-PKKFL(pThr)LGSK (HPLC-purified synthetic peptide)
Measured dissociation constant (10-6M): 5.4
Standard deviation of dissociation constant (10-6M): 0.3
Experiment method: competitive Fluorescence Polarization
Details of affinity fitting: Competitive binding equation in ProFit
Date of measurement: 2022.01.10
Number of measurements: 3
Measured pKd: 5.27