Details of experiments between P63104 and O43166
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: SIPA1L1
Protein B fragment: 1629-1638
Protein B site: 14-3-3_pThr1634
Protein B modification: (TPO)1634
Protein B sequence: RRPSYTLGMK
Protein B construct: RRPSY(pThr)LGMK (crude synthetic peptide)
Average holdup BI: 0.8
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): 8.91
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
Measured pKd: 4.47
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: SIPA1L1
Protein B fragment: 1559-1568
Protein B site: 14-3-3_pSer1564
Protein B modification: (SEP)1564
Protein B sequence: KFHALSSPQS
Protein B construct: KFHAL(pSer)SPQS (crude synthetic peptide)
Average holdup BI: 0.64
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): 5.51
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 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: SIPA1L1
Protein B fragment: 1399-1408
Protein B site: 14-3-3_pSer1404
Protein B modification: (SEP)1404
Protein B sequence: KSQAGSTPLT
Protein B construct: KSQAG(pSer)TPLT (crude synthetic peptide)
Average holdup BI: 0.22
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): 5.09
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: SIPA1L1
Protein B fragment: 1580-1589
Protein B site: 14-3-3_pSer1585
Protein B modification: (SEP)1585
Protein B sequence: LHRTLSDESI
Protein B construct: LHRTL(pSer)DESI (crude synthetic peptide)
Average holdup BI: 0.19
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): 1.89
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: 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: SIPA1L1
Protein B fragment: 1185-1194
Protein B site: 14-3-3_pThr1190
Protein B modification: (TPO)1190
Protein B sequence: IDRQNTQSDI
Protein B construct: IDRQN(pThr)QSDI (crude synthetic peptide)
Average holdup BI: -0.03
Holdup BI standard deviation: 0.06
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): 0.41
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: SIPA1L1
Protein B fragment: 1728-1737
Protein B site: 14-3-3_pSer1733
Protein B modification: (SEP)1733
Protein B sequence: SSSKDSSPTL
Protein B construct: SSSKD(pSer)SPTL (crude synthetic peptide)
Average holdup BI: -0.01
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): 0.39
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: SIPA1L1
Protein B fragment: 156-165
Protein B site: 14-3-3_pSer161
Protein B modification: (SEP)161
Protein B sequence: PEAYPSSPRK
Protein B construct: PEAYP(pSer)SPRK (crude synthetic peptide)
Average holdup BI: 0.01
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): 0.14
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: SIPA1L1
Protein B fragment: 1165-1174
Protein B site: 14-3-3_pSer1170
Protein B modification: (SEP)1170
Protein B sequence: TYRQKSMPEG
Protein B construct: TYRQK(pSer)MPEG (crude synthetic peptide)
Average holdup BI: 0.02
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): 0.1
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: SIPA1L1
Protein B fragment: 250-259
Protein B site: 14-3-3_pSer255
Protein B modification: (SEP)255
Protein B sequence: GGGKGSGFSL
Protein B construct: GGGKG(pSer)GFSL (crude synthetic peptide)
Average holdup BI: 0
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): 0.09
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.