Details of experiments between P63104 and O60343
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 699-708
Protein B site: 14-3-3_pSer704
Protein B modification: (SEP)704
Protein B sequence: LHTSFSAPSF
Protein B construct: LHTSF(pSer)APSF (crude synthetic peptide)
Measured dissociation constant (10-6M): <10
Immobilized partner concentration in holdup experiment (10-6M): 132
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: 1
Measured pKd: 5
Experiment series 2
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 563-572
Protein B site: 14-3-3_pThr568
Protein B modification: (TPO)568
Protein B sequence: AKRSLTSSLE
Protein B construct: AKRSL(pThr)SSLE (crude synthetic peptide)
Average holdup BI: 0.44
Holdup BI standard deviation: 0.03
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): 7.58
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: 3.77
Experiment series 3
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 592-601
Protein B site: 14-3-3_pSer597
Protein B modification: (SEP)597
Protein B sequence: FERSNSLASE
Protein B construct: FERSN(pSer)LASE (crude synthetic peptide)
Average holdup BI: 0.12
Holdup BI standard deviation: 0.02
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): 5.91
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: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 583-592
Protein B site: 14-3-3_pSer588
Protein B modification: (SEP)588
Protein B sequence: RGRLGSVDSF
Protein B construct: RGRLG(pSer)VDSF (crude synthetic peptide)
Average holdup BI: 0.36
Holdup BI standard deviation: 0.04
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): 5.29
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: 3.63
Experiment series 5
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 565-574
Protein B site: 14-3-3_pSer570
Protein B modification: (SEP)570
Protein B sequence: RSLTSSLENI
Protein B construct: RSLTS(pSer)LENI (crude synthetic peptide)
Average holdup BI: 0.12
Holdup BI standard deviation: 0.12
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): 1.11
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 6
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 595-604
Protein B site: 14-3-3_pSer600
Protein B modification: (SEP)600
Protein B sequence: SNSLASEKDY
Protein B construct: SNSLA(pSer)EKDY (crude synthetic peptide)
Average holdup BI: -0.03
Holdup BI standard deviation: 0.04
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): 0.85
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 7
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 480-489
Protein B site: 14-3-3_pSer485
Protein B modification: (SEP)485
Protein B sequence: IQRHLSSLTD
Protein B construct: IQRHL(pSer)SLTD (crude synthetic peptide)
Average holdup BI: 0.07
Holdup BI standard deviation: 0.11
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): 0.68
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 8
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 639-648
Protein B site: 14-3-3_pSer644
Protein B modification: (SEP)644
Protein B sequence: RAHTFSHPPS
Protein B construct: RAHTF(pSer)HPPS (crude synthetic peptide)
Average holdup BI: 0.1
Holdup BI standard deviation: 0.17
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): 0.59
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: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 336-345
Protein B site: 14-3-3_pSer341
Protein B modification: (SEP)341
Protein B sequence: RRRHASAPSH
Protein B construct: RRRHA(pSer)APSH (crude synthetic peptide)
Average holdup BI: 0.19
Holdup BI standard deviation: 0.54
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): 0.25
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
The affinity was below the detection threshold of the assay.
Experiment series 10
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 1202-1211
Protein B site: 14-3-3_pSer1207
Protein B modification: (SEP)1207
Protein B sequence: LERANSQLKR
Protein B construct: LERAN(pSer)QLKR (crude synthetic peptide)
Average holdup BI: 0.21
Holdup BI standard deviation: 0.91
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): 0.16
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 11
Protein A protein: YWHAZ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3zeta
Protein B protein: TBC1D4
Protein B fragment: 586-595
Protein B site: 14-3-3_pSer591
Protein B modification: (SEP)591
Protein B sequence: LGSVDSFERS
Protein B construct: LGSVD(pSer)FERS (crude synthetic peptide)
Average holdup BI: 0
Holdup BI standard deviation: 0.02
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): 0.16
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.