Details of experiments between P62258 and Q53ET0
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: CRTC2
Protein B fragment: 166-175
Protein B site: 14-3-3_pSer171
Protein B modification: (SEP)171
Protein B sequence: LNRTSSDSAL
Protein B construct: LNRTS(pSer)DSAL (crude synthetic peptide)
Average holdup BI: 0.22
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): 6.04
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.
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: CRTC2
Protein B fragment: 65-74
Protein B site: 14-3-3_pSer70
Protein B modification: (SEP)70
Protein B sequence: SHYGGSLPNV
Protein B construct: SHYGG(pSer)LPNV (crude synthetic peptide)
Average holdup BI: 0.47
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): 6
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: 3.8
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: CRTC2
Protein B fragment: 122-131
Protein B site: 14-3-3_pSer127
Protein B modification: (SEP)127
Protein B sequence: TRHIDSSPYS
Protein B construct: TRHID(pSer)SPYS (crude synthetic peptide)
Average holdup BI: 0.29
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): 4.7
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: 3.46
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: CRTC2
Protein B fragment: 122-131
Protein B site: 14-3-3_pSer127
Protein B modification: (SEP)127
Protein B sequence: TRHIDSSPYS
Protein B construct: TRHID(pSer)SPYS (crude synthetic peptide)
Average holdup BI: 0.18
Holdup BI standard deviation: 0.05
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): 2.81
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 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: CRTC2
Protein B fragment: 166-175
Protein B site: 14-3-3_pSer171
Protein B modification: (SEP)171
Protein B sequence: LNRTSSDSAL
Protein B construct: LNRTS(pSer)DSAL (crude synthetic peptide)
Average holdup BI: 0.22
Holdup BI standard deviation: 0.09
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): 2.62
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: 3.31
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: CRTC2
Protein B fragment: 65-74
Protein B site: 14-3-3_pSer70
Protein B modification: (SEP)70
Protein B sequence: SHYGGSLPNV
Protein B construct: SHYGG(pSer)LPNV (crude synthetic peptide)
Average holdup BI: 0.42
Holdup BI standard deviation: 0.17
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): 2.4
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 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: CRTC2
Protein B fragment: 487-496
Protein B site: 14-3-3_pSer492
Protein B modification: (SEP)492
Protein B sequence: PYSSPSLVLP
Protein B construct: PYSSP(pSer)LVLP (crude synthetic peptide)
Average holdup BI: -0.12
Holdup BI standard deviation: 0.17
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): 0.71
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: 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: CRTC2
Protein B fragment: 487-496
Protein B site: 14-3-3_pSer492
Protein B modification: (SEP)492
Protein B sequence: PYSSPSLVLP
Protein B construct: PYSSP(pSer)LVLP (crude synthetic peptide)
Average holdup BI: -0.21
Holdup BI standard deviation: 0.4
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): 0.62
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
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: CRTC2
Protein B fragment: 442-451
Protein B site: 14-3-3_pSer447
Protein B modification: (SEP)447
Protein B sequence: ADARRSQQQL
Protein B construct: ADARR(pSer)QQQL (crude synthetic peptide)
Average holdup BI: 0.25
Holdup BI standard deviation: 0.34
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): 0.44
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 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: CRTC2
Protein B fragment: 442-451
Protein B site: 14-3-3_pSer447
Protein B modification: (SEP)447
Protein B sequence: ADARRSQQQL
Protein B construct: ADARR(pSer)QQQL (crude synthetic peptide)
Average holdup BI: 0.06
Holdup BI standard deviation: 1.36
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.