Details of experiments between P27348 and Q53ET0
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
Protein B protein: CRTC2
Protein B fragment: 122-131
Protein B site: 14-3-3_pSer127
Protein B modification: (SEP)127
Protein B sequence: TRHIDSSPYS
Protein B construct: TRHID(pSer)SPYS (crude synthetic peptide)
Average holdup BI: 0.29
Holdup BI standard deviation: 0.04
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): 5.13
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: 3.49
Experiment series 2
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
Protein B protein: CRTC2
Protein B fragment: 166-175
Protein B site: 14-3-3_pSer171
Protein B modification: (SEP)171
Protein B sequence: LNRTSSDSAL
Protein B construct: LNRTS(pSer)DSAL (crude synthetic peptide)
Average holdup BI: 0.34
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): 3.68
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
Measured pKd: 3.6
Experiment series 3
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
Protein B protein: CRTC2
Protein B fragment: 65-74
Protein B site: 14-3-3_pSer70
Protein B modification: (SEP)70
Protein B sequence: SHYGGSLPNV
Protein B construct: SHYGG(pSer)LPNV (crude synthetic peptide)
Average holdup BI: 0.68
Holdup BI standard deviation: 0.36
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): 2.13
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: 4.21
Experiment series 4
Protein A protein: YWHAQ
Protein A fragment: 1-245
Protein A site: 14-3-3
Protein A construct: avi-his6-MBP-TEVsite-14-3-3tau
Protein B protein: CRTC2
Protein B fragment: 487-496
Protein B site: 14-3-3_pSer492
Protein B modification: (SEP)492
Protein B sequence: PYSSPSLVLP
Protein B construct: PYSSP(pSer)LVLP (crude synthetic peptide)
Average holdup BI: 0.03
Holdup BI standard deviation: 0.52
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): 0.04
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.