Details of experiments between P31946 and P27448
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: MARK3
Protein B fragment: 559-568
Protein B site: 14-3-3_pThr564
Protein B modification: (TPO)564
Protein B sequence: TASRSTFHGQ
Protein B construct: TASRS(pThr)FHGQ (crude synthetic peptide)
Measured dissociation constant (10-6M): <100
Immobilized partner concentration in holdup experiment (10-6M): 102
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: 1
Measured pKd: 4
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: MARK3
Protein B fragment: 596-605
Protein B site: 14-3-3_pSer601
Protein B modification: (SEP)601
Protein B sequence: TRSRGSTNLF
Protein B construct: TRSRG(pSer)TNLF (crude synthetic peptide)
Average holdup BI: 0.42
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): 7.45
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: 3.73
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: MARK3
Protein B fragment: 596-605
Protein B site: 14-3-3_pSer601
Protein B modification: (SEP)601
Protein B sequence: TRSRGSTNLF
Protein B construct: TRSRG(pSer)TNLF (crude synthetic peptide)
Average holdup BI: 0.38
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): 6.43
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: 3.78
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: MARK3
Protein B fragment: 91-100
Protein B site: 14-3-3_pSer96
Protein B modification: (SEP)96
Protein B sequence: QLNPTSLQKL
Protein B construct: QLNPT(pSer)LQKL (crude synthetic peptide)
Average holdup BI: 0.17
Holdup BI standard deviation: 0.09
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): 1.91
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 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: MARK3
Protein B fragment: 85-94
Protein B site: 14-3-3_pThr90
Protein B modification: (TPO)90
Protein B sequence: KIIDKTQLNP
Protein B construct: KIIDK(pThr)QLNP (crude synthetic peptide)
Average holdup BI: 0.2
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): 1.74
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: MARK3
Protein B fragment: 486-495
Protein B site: 14-3-3_pThr491
Protein B modification: (TPO)491
Protein B sequence: RKKSSTVPSS
Protein B construct: RKKSS(pThr)VPSS (crude synthetic peptide)
Average holdup BI: 0.07
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): 1.45
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
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: MARK3
Protein B fragment: 535-544
Protein B site: 14-3-3_pSer540
Protein B modification: (SEP)540
Protein B sequence: RTPVASTHSI
Protein B construct: RTPVA(pSer)THSI (crude synthetic peptide)
Average holdup BI: 0.06
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): 1.33
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: MARK3
Protein B fragment: 538-547
Protein B site: 14-3-3_pSer543
Protein B modification: (SEP)543
Protein B sequence: VASTHSISSA
Protein B construct: VASTH(pSer)ISSA (crude synthetic peptide)
Average holdup BI: -0.03
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): 0.78
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 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: MARK3
Protein B fragment: 91-100
Protein B site: 14-3-3_pSer96
Protein B modification: (SEP)96
Protein B sequence: QLNPTSLQKL
Protein B construct: QLNPT(pSer)LQKL (crude synthetic peptide)
Average holdup BI: 0.12
Holdup BI standard deviation: 0.17
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): 0.7
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: 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: MARK3
Protein B fragment: 486-495
Protein B site: 14-3-3_pThr491
Protein B modification: (TPO)491
Protein B sequence: RKKSSTVPSS
Protein B construct: RKKSS(pThr)VPSS (crude synthetic peptide)
Average holdup BI: 0.05
Holdup BI standard deviation: 0.08
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): 0.62
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 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: MARK3
Protein B fragment: 535-544
Protein B site: 14-3-3_pSer540
Protein B modification: (SEP)540
Protein B sequence: RTPVASTHSI
Protein B construct: RTPVA(pSer)THSI (crude synthetic peptide)
Average holdup BI: 0.03
Holdup BI standard deviation: 0.06
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): 0.5
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: 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: MARK3
Protein B fragment: 538-547
Protein B site: 14-3-3_pSer543
Protein B modification: (SEP)543
Protein B sequence: VASTHSISSA
Protein B construct: VASTH(pSer)ISSA (crude synthetic peptide)
Average holdup BI: -0.03
Holdup BI standard deviation: 0.07
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): 0.43
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: MARK3
Protein B fragment: 85-94
Protein B site: 14-3-3_pThr90
Protein B modification: (TPO)90
Protein B sequence: KIIDKTQLNP
Protein B construct: KIIDK(pThr)QLNP (crude synthetic peptide)
Average holdup BI: -0.04
Holdup BI standard deviation: 0.1
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): 0.39
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
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: MARK3
Protein B fragment: 206-215
Protein B site: 14-3-3_pThr211
Protein B modification: (TPO)211
Protein B sequence: GGKLDTFCGS
Protein B construct: GGKLD(pThr)FCGS (crude synthetic peptide)
Average holdup BI: -0.24
Holdup BI standard deviation: 1.88
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.