Details of experiments between the fragments Q4KWH8~1515-1524 and P27348~1-245
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: PLCH1
Protein A fragment: 1515-1524
Protein A site: 14-3-3_pThr1520
Protein A modification: (TPO)1520
Protein A sequence: SKSFVTTGIR
Protein A construct: SKSFV(pThr)TGIR (crude synthetic peptide)
Protein B protein: YWHAQ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3tau
Average holdup BI: 0.3
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): 3.52
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: PLCH1
Protein A fragment: 1515-1524
Protein A site: 14-3-3_pThr1520
Protein A modification: (TPO)1520
Protein A sequence: SKSFVTTGIR
Protein A construct: biotin-ttds-SKSFV(pThr)TGIR (HPLC-purified synthetic peptide)
Protein B protein: YWHAQ
Protein B fragment: 1-245
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3tau
Measured dissociation constant (10-6M): 401
Standard deviation of dissociation constant (10-6M): 26
Experiment method: competitive Fluorescence Polarization
Details of affinity fitting: Competitive binding equation in ProFit
Date of measurement: 2022.02.08
Number of measurements: 3
Measured pKd: 3.4