Details of experiments between P63104 and Q7Z309
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: FAM122B
Protein B fragment: 20-29
Protein B site: 14-3-3_pSer25
Protein B modification: (SEP)25
Protein B sequence: LRRSSSAPLI
Protein B construct: LRRSS(pSer)APLI (crude synthetic peptide)
Average holdup BI: 0.99
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): 7.53
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: 5.73
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: FAM122B
Protein B fragment: 28-37
Protein B site: 14-3-3_pSer33
Protein B modification: (SEP)33
Protein B sequence: LIHGLSDLSQ
Protein B construct: LIHGL(pSer)DLSQ (crude synthetic peptide)
Average holdup BI: -0.09
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): 1.58
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
The affinity was below the detection threshold of the assay.
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: FAM122B
Protein B fragment: 53-62
Protein B site: 14-3-3_pSer58
Protein B modification: (SEP)58
Protein B sequence: IMSRHSLEEG
Protein B construct: IMSRH(pSer)LEEG (crude synthetic peptide)
Average holdup BI: 0.05
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): 1.53
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: FAM122B
Protein B fragment: 116-125
Protein B site: 14-3-3_pThr121
Protein B modification: (TPO)121
Protein B sequence: PAPSPTRGFG
Protein B construct: PAPSP(pThr)RGFG (crude synthetic peptide)
Average holdup BI: 0.01
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): 0.18
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.
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: FAM122B
Protein B fragment: 45-54
Protein B site: 14-3-3_pSer50
Protein B modification: (SEP)50
Protein B sequence: RTRRNSTTIM
Protein B construct: RTRRN(pSer)TTIM (crude synthetic peptide)
Average holdup BI: -0.08
Holdup BI standard deviation: 0.34
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 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: FAM122B
Protein B fragment: 126-135
Protein B site: 14-3-3_pSer131
Protein B modification: (SEP)131
Protein B sequence: KMFVSSSGLP
Protein B construct: KMFVS(pSer)SGLP (crude synthetic peptide)
Average holdup BI: 0.12
Holdup BI standard deviation: 0.83
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.