Details of experiments between Q04917 and Q7Z401
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1586-1595
Protein B site: 14-3-3_pSer1591
Protein B modification: (SEP)1591
Protein B sequence: RSISTSGPLD
Protein B construct: RSIST(pSer)GPLD (crude synthetic peptide)
Average holdup BI: 0.3
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 7.73
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: 3.51
Experiment series 2
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1503-1512
Protein B site: 14-3-3_pSer1508
Protein B modification: (SEP)1508
Protein B sequence: RYFLKSSPST
Protein B construct: RYFLK(pSer)SPST (crude synthetic peptide)
Average holdup BI: 0.77
Holdup BI standard deviation: 0.07
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 6.17
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: 4.42
Experiment series 3
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1585-1594
Protein B site: 14-3-3_pThr1590
Protein B modification: (TPO)1590
Protein B sequence: ARSISTSGPL
Protein B construct: ARSIS(pThr)SGPL (crude synthetic peptide)
Average holdup BI: 0.53
Holdup BI standard deviation: 0.06
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 5.97
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: 3.93
Experiment series 4
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1030-1039
Protein B site: 14-3-3_pSer1035
Protein B modification: (SEP)1035
Protein B sequence: RNRNLSGGVL
Protein B construct: RNRNL(pSer)GGVL (crude synthetic peptide)
Average holdup BI: 0.37
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 4.87
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: 3.66
Experiment series 5
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1276-1285
Protein B site: 14-3-3_pSer1281
Protein B modification: (SEP)1281
Protein B sequence: RDRLWSSPAF
Protein B construct: RDRLW(pSer)SPAF (crude synthetic peptide)
Average holdup BI: 0.96
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 3.91
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: 5.22
Experiment series 6
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1146-1155
Protein B site: 14-3-3_pSer1151
Protein B modification: (SEP)1151
Protein B sequence: IQRMNSSFSV
Protein B construct: IQRMN(pSer)SFSV (crude synthetic peptide)
Average holdup BI: 0.22
Holdup BI standard deviation: 0.1
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 2.42
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
Measured pKd: 3.33
Experiment series 7
Protein A protein: YWHAH
Protein A fragment: 1-246
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3eta
Protein B protein: DENND4A
Protein B fragment: 1220-1229
Protein B site: 14-3-3_pSer1225
Protein B modification: (SEP)1225
Protein B sequence: TSRTPSIDLQ
Protein B construct: TSRTP(pSer)IDLQ (crude synthetic peptide)
Average holdup BI: -0.01
Holdup BI standard deviation: 0.07
Immobilized partner concentration in holdup experiment (10-6M): 130
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.17
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.