Details of experiments between P62258 and Q9NYF3
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 249-258
Protein B site: 14-3-3_pSer254
Protein B modification: (SEP)254
Protein B sequence: RSSPASSPEL
Protein B construct: RSSPA(pSer)SPEL (crude synthetic peptide)
Average holdup BI: 0.48
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 8.43
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: 3.81
Experiment series 2
Protein A protein: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 249-258
Protein B site: 14-3-3_pSer254
Protein B modification: (SEP)254
Protein B sequence: RSSPASSPEL
Protein B construct: RSSPA(pSer)SPEL (crude synthetic peptide)
Average holdup BI: 0.43
Holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 5.36
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: 3.81
Experiment series 3
Protein A protein: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 227-236
Protein B site: 14-3-3_pSer232
Protein B modification: (SEP)232
Protein B sequence: PQRRFSLSPS
Protein B construct: PQRRF(pSer)LSPS (crude synthetic peptide)
Average holdup BI: 0.15
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 4.51
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 4
Protein A protein: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 227-236
Protein B site: 14-3-3_pSer232
Protein B modification: (SEP)232
Protein B sequence: PQRRFSLSPS
Protein B construct: PQRRF(pSer)LSPS (crude synthetic peptide)
Average holdup BI: 0.21
Holdup BI standard deviation: 0.06
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 3.52
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: 3.27
Experiment series 5
Protein A protein: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 117-126
Protein B site: 14-3-3_pSer122
Protein B modification: (SEP)122
Protein B sequence: HCRSLSVPVD
Protein B construct: HCRSL(pSer)VPVD (crude synthetic peptide)
Average holdup BI: 0.82
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 2.45
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: 4.6
Experiment series 6
Protein A protein: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 294-303
Protein B site: 14-3-3_pSer299
Protein B modification: (SEP)299
Protein B sequence: RRLRPSLDFD
Protein B construct: RRLRP(pSer)LDFD (crude synthetic peptide)
Average holdup BI: -0.35
Holdup BI standard deviation: 0.97
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.34
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: YWHAE
Protein A fragment: 1-255
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Protein B protein: FAM53C
Protein B fragment: 294-303
Protein B site: 14-3-3_pSer299
Protein B modification: (SEP)299
Protein B sequence: RRLRPSLDFD
Protein B construct: RRLRP(pSer)LDFD (crude synthetic peptide)
Average holdup BI: 0.06
Holdup BI standard deviation: 0.29
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.16
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.