Details of experiments between O94915 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: FRYL
Protein A fragment: 1973-1982
Protein A site: 14-3-3_pSer1978
Protein A modification: (SEP)1978
Protein A sequence: LARTRSLSSL
Protein A construct: LARTR(pSer)LSSL (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: 0.89
Holdup BI standard deviation: 0.01
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): 9.24
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: 4.78
Experiment series 2
Protein A protein: FRYL
Protein A fragment: 1936-1945
Protein A site: 14-3-3_pSer1941
Protein A modification: (SEP)1941
Protein A sequence: NARSNSLRLS
Protein A construct: NARSN(pSer)LRLS (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: 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): 6.65
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: 3.72
Experiment series 3
Protein A protein: FRYL
Protein A fragment: 1954-1963
Protein A site: 14-3-3_pThr1959
Protein A modification: (TPO)1959
Protein A sequence: RRRSNTLDIM
Protein A construct: RRRSN(pThr)LDIM (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: 0.88
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): 5
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 4
Protein A protein: FRYL
Protein A fragment: 1940-1949
Protein A site: 14-3-3_pSer1945
Protein A modification: (SEP)1945
Protein A sequence: NSLRLSLIGD
Protein A construct: NSLRL(pSer)LIGD (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: -0.35
Holdup BI standard deviation: 0.53
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): 0.94
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: FRYL
Protein A fragment: 1930-1939
Protein A site: 14-3-3_pSer1935
Protein A modification: (SEP)1935
Protein A sequence: YLGYNSNARS
Protein A construct: YLGYN(pSer)NARS (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: 0.07
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 6
Protein A protein: FRYL
Protein A fragment: 1967-1976
Protein A site: 14-3-3_pSer1972
Protein A modification: (SEP)1972
Protein A sequence: INHSSSLART
Protein A construct: INHSS(pSer)LART (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: 0.07
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): 0.57
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: FRYL
Protein A fragment: 477-486
Protein A site: 14-3-3_pThr482
Protein A modification: (TPO)482
Protein A sequence: LPSGNTLRVK
Protein A construct: LPSGN(pThr)LRVK (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: 0.02
Holdup BI standard deviation: 0.1
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): 0.21
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: FRYL
Protein A fragment: 1909-1918
Protein A site: 14-3-3_pSer1914
Protein A modification: (SEP)1914
Protein A sequence: AANRKSTGQL
Protein A construct: AANRK(pSer)TGQL (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: -0.13
Holdup BI standard deviation: 0.94
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): 0.08
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: FRYL
Protein A fragment: 98-107
Protein A site: 14-3-3_pSer103
Protein A modification: (SEP)103
Protein A sequence: RSSTKSKGDE
Protein A construct: RSSTK(pSer)KGDE (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: 0.72
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): 0.04
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.