Details of experiments between P27348 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: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
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: 0.59
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 9.08
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
Measured pKd: 4.05
Experiment series 2
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
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: 0.56
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 7.15
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: 3.99
Experiment series 3
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
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: 0.03
Holdup BI standard deviation: 0.08
Immobilized partner concentration in holdup experiment (10-6M): 130
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 4
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
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): 130
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 5
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
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: 0.21
Immobilized partner concentration in holdup experiment (10-6M): 130
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.