Details of experiments between P63104 and Q9NSK0
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: KLC4
Protein B fragment: 585-594
Protein B site: 14-3-3_pSer590
Protein B modification: (SEP)590
Protein B sequence: MKRAASLNYL
Protein B construct: MKRAA(pSer)LNYL (crude synthetic peptide)
Average holdup BI: 0.83
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): 7.44
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: 4.56
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: KLC4
Protein B fragment: 549-558
Protein B site: 14-3-3_pSer554
Protein B modification: (SEP)554
Protein B sequence: LQRSGSLGKI
Protein B construct: LQRSG(pSer)LGKI (crude synthetic peptide)
Average holdup BI: 0.36
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): 5.31
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: 3.64
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: KLC4
Protein B fragment: 561-570
Protein B site: 14-3-3_pSer566
Protein B modification: (SEP)566
Protein B sequence: VLRRSSELLV
Protein B construct: VLRRS(pSer)ELLV (crude synthetic peptide)
Average holdup BI: 0.07
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): 0.96
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.