Details of experiments between P62258 and Q96PY6
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: NEK1
Protein B fragment: 409-418
Protein B site: 14-3-3_pSer414
Protein B modification: (SEP)414
Protein B sequence: WRNVLSAGGS
Protein B construct: WRNVL(pSer)AGGS (crude synthetic peptide)
Average holdup BI: 0.2
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): 3.27
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
Measured pKd: 3.24
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: NEK1
Protein B fragment: 409-418
Protein B site: 14-3-3_pSer414
Protein B modification: (SEP)414
Protein B sequence: WRNVLSAGGS
Protein B construct: WRNVL(pSer)AGGS (crude synthetic peptide)
Average holdup BI: 0.27
Holdup BI standard deviation: 0.07
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): 3.25
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: 3.49
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: NEK1
Protein B fragment: 280-289
Protein B site: 14-3-3_pSer285
Protein B modification: (SEP)285
Protein B sequence: FSKFGSQPIP
Protein B construct: FSKFG(pSer)QPIP (crude synthetic peptide)
Average holdup BI: 0.18
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): 2.4
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: 3.2
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: NEK1
Protein B fragment: 413-422
Protein B site: 14-3-3_pSer418
Protein B modification: (SEP)418
Protein B sequence: LSAGGSGEVK
Protein B construct: LSAGG(pSer)GEVK (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): 1.45
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
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: NEK1
Protein B fragment: 9-18
Protein B site: 14-3-3_pSer14
Protein B modification: (SEP)14
Protein B sequence: KIGEGSFGKA
Protein B construct: KIGEG(pSer)FGKA (crude synthetic peptide)
Average holdup BI: -0.04
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): 1.16
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 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: NEK1
Protein B fragment: 280-289
Protein B site: 14-3-3_pSer285
Protein B modification: (SEP)285
Protein B sequence: FSKFGSQPIP
Protein B construct: FSKFG(pSer)QPIP (crude synthetic peptide)
Average holdup BI: 0.14
Holdup BI standard deviation: 0.14
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): 0.98
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: NEK1
Protein B fragment: 136-145
Protein B site: 14-3-3_pThr141
Protein B modification: (TPO)141
Protein B sequence: LTKDGTVQLG
Protein B construct: LTKDG(pThr)VQLG (crude synthetic peptide)
Average holdup BI: 0.01
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): 0.75
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: NEK1
Protein B fragment: 151-160
Protein B site: 14-3-3_pThr156
Protein B modification: (TPO)156
Protein B sequence: RVLNSTVELA
Protein B construct: RVLNS(pThr)VELA (crude synthetic peptide)
Average holdup BI: -0.03
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): 0.52
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: NEK1
Protein B fragment: 1047-1056
Protein B site: 14-3-3_pSer1052
Protein B modification: (SEP)1052
Protein B sequence: MLRTCSLPDL
Protein B construct: MLRTC(pSer)LPDL (crude synthetic peptide)
Average holdup BI: -0.27
Holdup BI standard deviation: 0.62
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): 0.38
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: NEK1
Protein B fragment: 294-303
Protein B site: 14-3-3_pSer299
Protein B modification: (SEP)299
Protein B sequence: ASGQNSISVM
Protein B construct: ASGQN(pSer)ISVM (crude synthetic peptide)
Average holdup BI: 0.19
Holdup BI standard deviation: 0.52
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): 0.28
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 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: NEK1
Protein B fragment: 9-18
Protein B site: 14-3-3_pSer14
Protein B modification: (SEP)14
Protein B sequence: KIGEGSFGKA
Protein B construct: KIGEG(pSer)FGKA (crude synthetic peptide)
Average holdup BI: -0.01
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): 0.22
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 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: NEK1
Protein B fragment: 136-145
Protein B site: 14-3-3_pThr141
Protein B modification: (TPO)141
Protein B sequence: LTKDGTVQLG
Protein B construct: LTKDG(pThr)VQLG (crude synthetic peptide)
Average holdup BI: 0.01
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): 0.18
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 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: NEK1
Protein B fragment: 1121-1130
Protein B site: 14-3-3_pSer1126
Protein B modification: (SEP)1126
Protein B sequence: PGEEYSEEEE
Protein B construct: PGEEY(pSer)EEEE (crude synthetic peptide)
Average holdup BI: -0.26
Holdup BI standard deviation: 1.63
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): 0.15
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 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: NEK1
Protein B fragment: 151-160
Protein B site: 14-3-3_pThr156
Protein B modification: (TPO)156
Protein B sequence: RVLNSTVELA
Protein B construct: RVLNS(pThr)VELA (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.07
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 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: NEK1
Protein B fragment: 413-422
Protein B site: 14-3-3_pSer418
Protein B modification: (SEP)418
Protein B sequence: LSAGGSGEVK
Protein B construct: LSAGG(pSer)GEVK (crude synthetic peptide)
Average holdup BI: 0
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): 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.