Details of experiments between P31946 and Q96FS4
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: SIPA1
Protein B fragment: 903-912
Protein B site: 14-3-3_pSer908
Protein B modification: (SEP)908
Protein B sequence: LPRTLSLRNS
Protein B construct: LPRTL(pSer)LRNS (crude synthetic peptide)
Average holdup BI: 0.47
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): 5.62
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
Measured pKd: 3.94
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: SIPA1
Protein B fragment: 903-912
Protein B site: 14-3-3_pSer908
Protein B modification: (SEP)908
Protein B sequence: LPRTLSLRNS
Protein B construct: LPRTL(pSer)LRNS (crude synthetic peptide)
Average holdup BI: 0.54
Holdup BI standard deviation: 0.08
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): 3.88
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: 3.95
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: SIPA1
Protein B fragment: 50-59
Protein B site: 14-3-3_pSer55
Protein B modification: (SEP)55
Protein B sequence: LLRSGSDAGE
Protein B construct: LLRSG(pSer)DAGE (crude synthetic peptide)
Average holdup BI: -0.14
Holdup BI standard deviation: 0.25
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): 0.73
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: 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: SIPA1
Protein B fragment: 554-563
Protein B site: 14-3-3_pSer559
Protein B modification: (SEP)559
Protein B sequence: RFGLPSLGGR
Protein B construct: RFGLP(pSer)LGGR (crude synthetic peptide)
Average holdup BI: -0.15
Holdup BI standard deviation: 0.31
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 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: SIPA1
Protein B fragment: 832-841
Protein B site: 14-3-3_pSer837
Protein B modification: (SEP)837
Protein B sequence: LHSQNSLSPR
Protein B construct: LHSQN(pSer)LSPR (crude synthetic peptide)
Average holdup BI: 0.05
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): 0.43
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
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: SIPA1
Protein B fragment: 554-563
Protein B site: 14-3-3_pSer559
Protein B modification: (SEP)559
Protein B sequence: RFGLPSLGGR
Protein B construct: RFGLP(pSer)LGGR (crude synthetic peptide)
Average holdup BI: 0.23
Holdup BI standard deviation: 0.55
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): 0.34
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: 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: SIPA1
Protein B fragment: 50-59
Protein B site: 14-3-3_pSer55
Protein B modification: (SEP)55
Protein B sequence: LLRSGSDAGE
Protein B construct: LLRSG(pSer)DAGE (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.03
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.
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: SIPA1
Protein B fragment: 832-841
Protein B site: 14-3-3_pSer837
Protein B modification: (SEP)837
Protein B sequence: LHSQNSLSPR
Protein B construct: LHSQN(pSer)LSPR (crude synthetic peptide)
Average holdup BI: 0.02
Holdup BI standard deviation: 0.14
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.
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: SIPA1
Protein B fragment: 675-684
Protein B site: 14-3-3_pSer680
Protein B modification: (SEP)680
Protein B sequence: RLQLVSRGCE
Protein B construct: RLQLV(pSer)RGCE (crude synthetic peptide)
Average holdup BI: -0.01
Holdup BI standard deviation: 0.06
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): 0.04
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.