Details of experiments between P31946 and Q96NE9
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: FRMD6
Protein B fragment: 517-526
Protein B site: 14-3-3_pSer522
Protein B modification: (SEP)522
Protein B sequence: KLRGQSTDSL
Protein B construct: KLRGQ(pSer)TDSL (crude synthetic peptide)
Average holdup BI: 0.22
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): 5.55
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 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: FRMD6
Protein B fragment: 520-529
Protein B site: 14-3-3_pSer525
Protein B modification: (SEP)525
Protein B sequence: GQSTDSLPQT
Protein B construct: GQSTD(pSer)LPQT (crude synthetic peptide)
Average holdup BI: 0.21
Holdup BI standard deviation: 0.04
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): 3.77
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 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: FRMD6
Protein B fragment: 539-548
Protein B site: 14-3-3_pSer544
Protein B modification: (SEP)544
Protein B sequence: DRHSLSLDDI
Protein B construct: DRHSL(pSer)LDDI (crude synthetic peptide)
Average holdup BI: 0.31
Holdup BI standard deviation: 0.13
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): 2.52
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: 3.65
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: FRMD6
Protein B fragment: 517-526
Protein B site: 14-3-3_pSer522
Protein B modification: (SEP)522
Protein B sequence: KLRGQSTDSL
Protein B construct: KLRGQ(pSer)TDSL (crude synthetic peptide)
Average holdup BI: 0.16
Holdup BI standard deviation: 0.13
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): 1.33
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: FRMD6
Protein B fragment: 520-529
Protein B site: 14-3-3_pSer525
Protein B modification: (SEP)525
Protein B sequence: GQSTDSLPQT
Protein B construct: GQSTD(pSer)LPQT (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.15
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): 0.01
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.