Details of experiments between P31946 and O60333
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: KIF1B
Protein B fragment: 1048-1057
Protein B site: 14-3-3_pSer1053
Protein B modification: (SEP)1053
Protein B sequence: RWRSNSLNNG
Protein B construct: RWRSN(pSer)LNNG (crude synthetic peptide)
Average holdup BI: 0.8
Holdup BI standard deviation: 0.03
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): 7.25
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: 4.6
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: KIF1B
Protein B fragment: 1136-1145
Protein B site: 14-3-3_pSer1141
Protein B modification: (SEP)1141
Protein B sequence: MRRQFSAPNL
Protein B construct: MRRQF(pSer)APNL (crude synthetic peptide)
Average holdup BI: 0.94
Holdup BI standard deviation: 0.01
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): 6.18
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
Measured pKd: 5.18
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: KIF1B
Protein B fragment: 1048-1057
Protein B site: 14-3-3_pSer1053
Protein B modification: (SEP)1053
Protein B sequence: RWRSNSLNNG
Protein B construct: RWRSN(pSer)LNNG (crude synthetic peptide)
Average holdup BI: 0.84
Holdup BI standard deviation: 0.11
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): 5.11
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: 4.58
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: KIF1B
Protein B fragment: 1136-1145
Protein B site: 14-3-3_pSer1141
Protein B modification: (SEP)1141
Protein B sequence: MRRQFSAPNL
Protein B construct: MRRQF(pSer)APNL (crude synthetic peptide)
Average holdup BI: 0.98
Holdup BI standard deviation: 0.02
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): 2.8
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: 5.65
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: KIF1B
Protein B fragment: 887-896
Protein B site: 14-3-3_pSer892
Protein B modification: (SEP)892
Protein B sequence: FKLVGSSPIF
Protein B construct: FKLVG(pSer)SPIF (crude synthetic peptide)
Average holdup BI: 0.87
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): 2.76
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: 4.7
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: KIF1B
Protein B fragment: 887-896
Protein B site: 14-3-3_pSer892
Protein B modification: (SEP)892
Protein B sequence: FKLVGSSPIF
Protein B construct: FKLVG(pSer)SPIF (crude synthetic peptide)
Average holdup BI: 0.85
Holdup BI standard deviation: 0.08
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): 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: 4.74
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: KIF1B
Protein B fragment: 1052-1061
Protein B site: 14-3-3_pSer1057
Protein B modification: (SEP)1057
Protein B sequence: TRSGLSLEEL
Protein B construct: TRSGL(pSer)LEEL (crude synthetic peptide)
Average holdup BI: 0.13
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): 2.18
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: KIF1B
Protein B fragment: 517-526
Protein B site: 14-3-3_pThr522
Protein B modification: (TPO)522
Protein B sequence: REDGGTLGVF
Protein B construct: REDGG(pThr)LGVF (crude synthetic peptide)
Average holdup BI: 0.07
Holdup BI standard deviation: 0.04
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): 1.65
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: KIF1B
Protein B fragment: 1052-1061
Protein B site: 14-3-3_pSer1057
Protein B modification: (SEP)1057
Protein B sequence: TRSGLSLEEL
Protein B construct: TRSGL(pSer)LEEL (crude synthetic peptide)
Average holdup BI: 0.18
Holdup BI standard deviation: 0.13
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): 1.56
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: 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: KIF1B
Protein B fragment: 1482-1491
Protein B site: 14-3-3_pSer1487
Protein B modification: (SEP)1487
Protein B sequence: RPRGDSLILE
Protein B construct: RPRGD(pSer)LILE (crude synthetic peptide)
Average holdup BI: 0.06
Holdup BI standard deviation: 0.05
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): 1.47
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: 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: KIF1B
Protein B fragment: 658-667
Protein B site: 14-3-3_pSer663
Protein B modification: (SEP)663
Protein B sequence: RLDADSDSGD
Protein B construct: RLDAD(pSer)DSGD (crude synthetic peptide)
Average holdup BI: -0.18
Holdup BI standard deviation: 0.17
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): 1.33
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: 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: KIF1B
Protein B fragment: 517-526
Protein B site: 14-3-3_pThr522
Protein B modification: (TPO)522
Protein B sequence: REDGGTLGVF
Protein B construct: REDGG(pThr)LGVF (crude synthetic peptide)
Average holdup BI: 0.12
Holdup BI standard deviation: 0.16
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): 0.76
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: 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: KIF1B
Protein B fragment: 1482-1491
Protein B site: 14-3-3_pSer1487
Protein B modification: (SEP)1487
Protein B sequence: RPRGDSLILE
Protein B construct: RPRGD(pSer)LILE (crude synthetic peptide)
Average holdup BI: 0.03
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): 0.46
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.