Details of experiments between P62258 and O60292
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: SIPA1L3
Protein B fragment: 1377-1386
Protein B site: 14-3-3_pSer1382
Protein B modification: (SEP)1382
Protein B sequence: RPKLYSSGSS
Protein B construct: biotin-ttds-RPKLY(pSer)SGSS (HPLC-purified synthetic peptide)
Measured dissociation constant (10-6M): below threshold
Experiment method: competitive Fluorescence Polarization
Details of affinity fitting: Competitive binding equation in ProFit
Date of measurement: 2022.02.09
Number of measurements: 3
The affinity was below the detection threshold of the assay.
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: SIPA1L3
Protein B fragment: 1130-1139
Protein B site: 14-3-3_pSer1135
Protein B modification: (SEP)1135
Protein B sequence: SKRPVSFPET
Protein B construct: SKRPV(pSer)FPET (crude synthetic peptide)
Average holdup BI: 0.48
Holdup BI standard deviation: 0.03
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): 6.93
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: 3.82
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: SIPA1L3
Protein B fragment: 386-395
Protein B site: 14-3-3_pSer391
Protein B modification: (SEP)391
Protein B sequence: ASRAHSLGGL
Protein B construct: ASRAH(pSer)LGGL (crude synthetic peptide)
Average holdup BI: 0.49
Holdup BI standard deviation: 0.03
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): 6.87
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: 3.92
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: SIPA1L3
Protein B fragment: 1468-1477
Protein B site: 14-3-3_pSer1473
Protein B modification: (SEP)1473
Protein B sequence: RPLPFSDPKK
Protein B construct: RPLPF(pSer)DPKK (crude synthetic peptide)
Average holdup BI: 0.43
Holdup BI standard deviation: 0.03
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): 6.41
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: 3.82
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: SIPA1L3
Protein B fragment: 1130-1139
Protein B site: 14-3-3_pSer1135
Protein B modification: (SEP)1135
Protein B sequence: SKRPVSFPET
Protein B construct: SKRPV(pSer)FPET (crude synthetic peptide)
Average holdup BI: 0.43
Holdup BI standard deviation: 0.03
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): 6.32
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 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: SIPA1L3
Protein B fragment: 386-395
Protein B site: 14-3-3_pSer391
Protein B modification: (SEP)391
Protein B sequence: ASRAHSLGGL
Protein B construct: ASRAH(pSer)LGGL (crude synthetic peptide)
Average holdup BI: 0.5
Holdup BI standard deviation: 0.05
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): 5.78
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: 3.84
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: SIPA1L3
Protein B fragment: 1468-1477
Protein B site: 14-3-3_pSer1473
Protein B modification: (SEP)1473
Protein B sequence: RPLPFSDPKK
Protein B construct: RPLPF(pSer)DPKK (crude synthetic peptide)
Average holdup BI: 0.39
Holdup BI standard deviation: 0.08
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): 4.49
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: 3.66
Experiment series 8
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: SIPA1L3
Protein B fragment: 1377-1386
Protein B site: 14-3-3_pSer1382
Protein B modification: (SEP)1382
Protein B sequence: RPKLYSSGSS
Protein B construct: RPKLY(pSer)SGSS (crude synthetic peptide)
Average holdup BI: 0.06
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): 2.63
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: 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: SIPA1L3
Protein B fragment: 1377-1386
Protein B site: 14-3-3_pSer1382
Protein B modification: (SEP)1382
Protein B sequence: RPKLYSSGSS
Protein B construct: RPKLY(pSer)SGSS (crude synthetic peptide)
Average holdup BI: 0.12
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): 2.27
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 10
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: SIPA1L3
Protein B fragment: 167-176
Protein B site: 14-3-3_pSer172
Protein B modification: (SEP)172
Protein B sequence: RHRSSSEITL
Protein B construct: RHRSS(pSer)EITL (crude synthetic peptide)
Average holdup BI: 0.21
Holdup BI standard deviation: 0.1
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): 2.17
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
Measured pKd: 3.26
Experiment series 11
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: SIPA1L3
Protein B fragment: 167-176
Protein B site: 14-3-3_pSer172
Protein B modification: (SEP)172
Protein B sequence: RHRSSSEITL
Protein B construct: RHRSS(pSer)EITL (crude synthetic peptide)
Average holdup BI: 0.18
Holdup BI standard deviation: 0.13
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): 1.35
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 12
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: SIPA1L3
Protein B fragment: 92-101
Protein B site: 14-3-3_pSer97
Protein B modification: (SEP)97
Protein B sequence: ALREHSNPSP
Protein B construct: ALREH(pSer)NPSP (crude synthetic peptide)
Average holdup BI: -0.24
Holdup BI standard deviation: 0.21
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): 1.35
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 13
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: SIPA1L3
Protein B fragment: 1539-1548
Protein B site: 14-3-3_pSer1544
Protein B modification: (SEP)1544
Protein B sequence: LQRTLSDESL
Protein B construct: LQRTL(pSer)DESL (crude synthetic peptide)
Average holdup BI: 0.08
Holdup BI standard deviation: 0.06
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): 1.34
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 14
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: SIPA1L3
Protein B fragment: 92-101
Protein B site: 14-3-3_pSer97
Protein B modification: (SEP)97
Protein B sequence: ALREHSNPSP
Protein B construct: ALREH(pSer)NPSP (crude synthetic peptide)
Average holdup BI: -0.12
Holdup BI standard deviation: 0.21
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): 0.54
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 15
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: SIPA1L3
Protein B fragment: 1120-1129
Protein B site: 14-3-3_pSer1125
Protein B modification: (SEP)1125
Protein B sequence: VPFRESQPLH
Protein B construct: VPFRE(pSer)QPLH (crude synthetic peptide)
Average holdup BI: -0.01
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): 0.1
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 16
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: SIPA1L3
Protein B fragment: 1539-1548
Protein B site: 14-3-3_pSer1544
Protein B modification: (SEP)1544
Protein B sequence: LQRTLSDESL
Protein B construct: LQRTL(pSer)DESL (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.03
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): 0.03
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 17
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: SIPA1L3
Protein B fragment: 1120-1129
Protein B site: 14-3-3_pSer1125
Protein B modification: (SEP)1125
Protein B sequence: VPFRESQPLH
Protein B construct: VPFRE(pSer)QPLH (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.11
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): 0.02
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.