Details of experiments between P31946 and Q6NUK4
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: REEP3
Protein B fragment: 237-246
Protein B site: 14-3-3_pSer242
Protein B modification: (SEP)242
Protein B sequence: EVRYGSLKYK
Protein B construct: EVRYG(pSer)LKYK (crude synthetic peptide)
Average holdup BI: 0.46
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): 5.49
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: 3.8
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: REEP3
Protein B fragment: 237-246
Protein B site: 14-3-3_pSer242
Protein B modification: (SEP)242
Protein B sequence: EVRYGSLKYK
Protein B construct: EVRYG(pSer)LKYK (crude synthetic peptide)
Average holdup BI: 0.42
Holdup BI standard deviation: 0.09
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): 4.08
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: 3.85
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: REEP3
Protein B fragment: 145-154
Protein B site: 14-3-3_pSer150
Protein B modification: (SEP)150
Protein B sequence: TERLRSFSMH
Protein B construct: TERLR(pSer)FSMH (crude synthetic peptide)
Average holdup BI: 0.45
Holdup BI standard deviation: 0.3
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): 1.49
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: 3.78
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: REEP3
Protein B fragment: 145-154
Protein B site: 14-3-3_pSer150
Protein B modification: (SEP)150
Protein B sequence: TERLRSFSMH
Protein B construct: TERLR(pSer)FSMH (crude synthetic peptide)
Average holdup BI: 0.09
Holdup BI standard deviation: 0.11
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): 0.83
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 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: REEP3
Protein B fragment: 205-214
Protein B site: 14-3-3_pSer210
Protein B modification: (SEP)210
Protein B sequence: AEGPYSDNEM
Protein B construct: AEGPY(pSer)DNEM (crude synthetic peptide)
Average holdup BI: 0.11
Holdup BI standard deviation: 0.5
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): 0.17
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.