Details of experiments between Q86TI0 and P63104
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: TBC1D1
Protein A fragment: 232-241
Protein A site: 14-3-3_pSer237
Protein A modification: (SEP)237
Protein A sequence: MRKSFSQPGL
Protein A construct: MRKSF(pSer)QPGL (crude synthetic peptide)
Protein B protein: YWHAZ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.64
Holdup BI standard deviation: 0.13
Immobilized partner concentration in holdup experiment (10-6M): 132
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 2.47
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: 4.12
Experiment series 2
Protein A protein: TBC1D1
Protein A fragment: 622-631
Protein A site: 14-3-3_pSer627
Protein A modification: (SEP)627
Protein A sequence: LMRYHSVSTE
Protein A construct: LMRYH(pSer)VSTE (crude synthetic peptide)
Protein B protein: YWHAZ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.1
Holdup BI standard deviation: 0.08
Immobilized partner concentration in holdup experiment (10-6M): 132
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 1.32
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 3
Protein A protein: TBC1D1
Protein A fragment: 561-570
Protein A site: 14-3-3_pSer566
Protein A modification: (SEP)566
Protein A sequence: KLLGSSEDLS
Protein A construct: KLLGS(pSer)EDLS (crude synthetic peptide)
Protein B protein: YWHAZ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.04
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 132
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: 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: TBC1D1
Protein A fragment: 591-600
Protein A site: 14-3-3_pThr596
Protein A modification: (TPO)596
Protein A sequence: RRRANTLSHF
Protein A construct: RRRAN(pThr)LSHF (crude synthetic peptide)
Protein B protein: YWHAZ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: -0.24
Holdup BI standard deviation: 0.72
Immobilized partner concentration in holdup experiment (10-6M): 132
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.42
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: TBC1D1
Protein A fragment: 566-575
Protein A site: 14-3-3_pSer571
Protein A modification: (SEP)571
Protein A sequence: SEDLSSDSES
Protein A construct: SEDLS(pSer)DSES (crude synthetic peptide)
Protein B protein: YWHAZ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: -0.41
Holdup BI standard deviation: 2.27
Immobilized partner concentration in holdup experiment (10-6M): 132
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.